Applied Polymer Light Microscopy

APPLIED POLYMER LIGHT MICROSCOPY

APPLIED POLYMER LIGHT MICROSCOPY

Edited by

D. A. HEMSLEY

Polymer Microscopy Services. Loughborough. UK

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© 1989 ELSEVIER SCIENCE PUBLISHERS LTD SOFTCOVER REPRINT OF THE HARDCOVER 1ST EDITION 1989 British Library Cataloguing in Publication Data

Applied polymer light microscopy I. Polymers. Microscopy I. Hemsley, D. A (Derek A) 547.7'046

ISBN-13: 978-94-011-7476-3 e-ISBN-13: 978-94-011-7474-9 DOl: 10.1007/978-94-011-7474-9

Library of Congress Cataloging in Publication Data

Applied polymer light microscopy/edited by D. A Hemsley. p. cm.

Bibliography: p. Includes index. ISBN-13: 978-94-011-7476-3 I. Polymers-Optical properties. 2. Polymers-Surfaces. 3. Microscope and microscopy-Technique. I. Hemsley, D. A (Derek A) QD381.9.066A67 1989 547.7'046-dcI9

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Preface

Synthetic polymers make excellent specimens for light microscopy. Despite this, the use of the technique, at least in its advanced forms, is not so widespread as might be expected. Although reliable and relevant data are difficult to find and quantify, it seems that in other fields of materials science and technology there is a greater readiness to tum to the microscope in research, in industrial problem solving, or for quality assessment and control. It also seems that the reasons for the present situation are partly historical, partly the result of the structure of the plastics and rubber industries, and partly the education and training background of senior staff who tend to be chemistry or engineering based. In neither field does light microscopy feature strongly in the basic training.

The primary aim of this book is to provide some insight into the range oflight microscopy techniques applicable to polymeric specimens, and to highlight typical applications to commercial polymers and polymer products. Where appropriate, the optical techniques involved are discussed in some detail. However, it has not been the intention to produce a light microscopy textbook dealing with the principles and design of the basic instrument. Many such texts are available, and selected examples are cited in the reference list at the end of most chapters.

Light microscopy is but a part of the broad field of polymer microscopy. The chart outlines this field and shows, in block capitals, the subject areas with which this book is specifically concerned. The chart is by no means complete. Electron microscopy could itself be subdivided into a set of more specific techniques, as could microradiography and acoustic microscopy. Furthermore, certain light microscopy

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Preface vii

methods, such as scanning optical techniques, have also been omitted on the basis that, although potentially valuable, they have not as yet established themselves for general and routine polymer work.

Chapter I describes the principles and practice of specimen preparation for light microscopy. Good specimen preparation is a prerequisite of good microscopy; it is difficult to overemphasize that time and care spent at this stage of a microscopical examination will be well rewarded.

Chapter 2 looks at aspects of some image formation in the basic light microscope and at phase contrast microscopy. As in all chapters, examples are given of typical application areas.

Chapters 3 and 4 are concerned with what is traditionally seen as the major technique in the light microscopy of plastics. Polarized light methods may be applied in a qualitative or quantitative manner; a chapter is devoted to each. The quantitative methods described in Chapter 4 need to be employed with care. The optical measurements themselves present few problems. Their interpretation is more difficult but they can nevertheless provide valuable data about the organization of the molecules in manufactured products, and the magnitude, type and direction of frozen-in stresses.

Chapter 5 covers two techniques related by a similarity in the type of image produced. Both are essentially contrast enhancing methods and as such compete with phase contrast. Each technique has its own particular advantages and disadvantages; an objective of this section of the book is to help in the selection of the most appropriate technique for a particular combination of specimen type and information requirement.

The broad subject of interference microscopy is discussed in Chapter 6. The potential for the techniques discussed is considerable and has yet to be fully realized in the examination of polymer products and systems. All the methods discussed are quantitative, although some may also be used to advantage in a qualitative mode. The microscopy of composites, blends and multilayer polymer structures often involves the identification of phases in the material; transmitted light microinterferometry can be of assistance in this task. Reflected light methods have been applied to polymer surfaces; special emphasis is given to this application in Chapter 6.

Chapter 7 takes microscopy outside the visible spectrum and looks at the use of ultraviolet radiation in polymer work. Here again both qualitative and quantitative methods are involved. Two basic techniques are discusssed; in one the image is formed by the ultraviolet radiation

viii Preface

itself, and in the other the radiation is used to promote fluorescence and the image is viewed using visible light. The latter perhaps provides the justification for inclusion of these UV methods amongst the visible light techniques.

One of the fundamental difficulties of polymer microscopy is image interpretation. Whilst the practical use of the microscope can be readily demonstrated, interpretation of the image it produces involves an understanding both of the characteristics of the optical system and of the optical properties of the specimen. This book is intended to help with at least the first of these requirements. Interpretation is easier the greater the amount of information available. For all but the simplest specimens it is therefore advisable to employ more than one of the techniques described to establish as complete a picture of its microstructure as possible. The 'quick look' using a single technique can easily be the route to an interpretational disaster!

D. A. HEMSLEY

Contents

~eface v

List of Contributors xii

1 Specimen Preparation A. D. CURSON 1.1 Introduction 1 1.2 Initial Approach to Preparing the Specimen 3 1.3 Area of the Specimen . 4 1.4 Surfaces 5 1.5 Sectioning (Sectional Slices) 18 1.6 Melt Pressings 36 1.7 Staining 36 1.8 Final Comments. 37

2 Basic Light Microscopy and the Phase Contrast Microscope D. A. HEMSLEY

2.1 Introduction . 39 2.2 Synthetic Polymers as Specimens for Light Microscopy 40 2.3 Light versus Electron Microscopy of Polymers 43 2.4 Basic Light Microscopy 46 2.5 Applications for Common Light Microscopy 52 2.6 Phase Contrast Microscopy . 60 2.7 Applications for Phase Contrast Microscopy 65 2.8 Dark Ground Microscopy 70

References 71

3 Polarized Light: Theory and Measurements B. P. SAVILLE 3.1 Introduction 73 3.2 Light and Its Interaction with Matter. 73 3.3 Elliptically and Circularly Polarized Light 81

ix

x Contents

3.4 The Uniaxial Indicatrix 82 3.5 The Biaxial Indicatrix 83 3.6 Methods of Producing Polarized Light 85 3.7 Types of Birefringence 87 3.8 The Passage of Polarized Light through Thin Birefringent

Plates . 88 3.9 Polarization Colours . 91 3.10 Relation between Orientation and Birefringence 93 3.11 The Polarizing Microscope 96 3.12 Measurement of Optical Path Difference 99 3.13 Compensators 100 3.14 Dispersion of Birefringence 104 3.15 Spectrophotometric Method 105 3.16 The Wedge Method 107 3.17 Use of the Abbe Refractometer 108

References . 108 Bibliography 109

4 Polarized Light: Qualitative Microscopy 4.1 Introduction .

B. P. SAVILLE III 112 117 125 132 136 144 145 149

4.2 Spherulites 4.3 Theory of Spherulitic Crystallization 4.4 Different Types of Spherulite 4.5 Spherulitic Forms of Polypropylene 4.6 Effect of Processing on Spherulites 4.7 Small Angle Light Scattering 4.8 Molecular Orientation

References

5 Modulation Contrast and Differential Interference Contrast Techniques R. HOFFMAN

5.1 Introduction. 151 5.2 General Principles 151 5.3 The Modulation Contrast System. 155 5.4 Differential Interference Contrast 162 5.5 Adjusting the Contrast Systems 167 5.6 Comparison with the Phase Contrast Microscope 169 5.7 Reflected Light Work 169 5.8 Image Interpretation 170 5.9 Applications to Polymers 173

References 183

6 Interference Microscopy of Polymers D. A. HEMSLEY 6.1 Introduction . 185 6.2 The Basic Principles of Quantitative Microinterferometry 186 6.3 Reflected Light Applications 192

Contents xi

6.4 Interpretation of Surface Interferograms 201 6.5 Some Reflected Light Systems Applicable to Polymers 207 6.6 Transmitted Light Interference Microscopy. 212 6.7 Transmitted Light Systems 222 6.8 Compensators 230

References 231

7 Ultraviolet and Fluorescence Microscopy P. CALVERT and N. C. BILLINGHAM

7.1 Introduction . 233 7.2 Equipment and Techniques. 235 7.3 Applications to Non-polymer Materials 242 7.4 Non-Microscopic Applications of Fluorescence from Polymers 244 7.5 Applications ofUV Microscopy to Synthetic Polymers 245

References 270

Index. 273

List of Contributors

N. C. BILLINGHAM

School of Chemistry and Molecular Science, University of Sussex, Brighton BNl 9RH, UK

P. CALVERT

School of Chemistry and Molecular Science, University of Sussex, Brighton BNl 9RH, UK Present address: Arizona Materials Laboratories, University of Arizona, Tucson, Arizona, USA

A. D. CURSON

Materials Science Group, ICI Advanced Materials, Wilton Materials Research Centre, PO Box 90, Wilton, Middlesbrough, Cleveland TS6 BJE, UK

D. A. HEMSLEY

Polymer Microscopy Services, 52 Springfield Close, Burton on the Wolds, Loughborough, Leicestershire LE12 5AN, UK

R. HOFFMAN

Modulation Optics, 100 Forest Drive, Greenvale, New York 11548, USA

B. P . SAVILLE

Institute of Polymer Technology, University of Technology, Loughborough, Leicestershire LEI 1 3TV, UK Present address: Department of Textiles, The Polytechnic, Queensgate, Huddersfield, West Yorkshire HDI 3DH, UK

xii

1

Specimen Preparation

A. D. CURSON Materials Science Group, ICI Advanced Materials,

Wilton Materials Research Centre, Middlesbrough, UK

1.1 INTRODUCTION

Specimen preparation is the gateway to good and meaningful microscopy. Given a well prepared specimen an experienced microscopist will obtain the maximum amount of accurate information; a poor microscopist will generate only poor resultS: With a poorly prepared specimen even a good microscopist, equipped with the best microscope, can expect only poor results. The effect of poor specimen preparation can variously destroy the structure, modify its appearance, or even introduce a totally new structure into the specimen being examined. The experienced microscopist will recognise artefacts of preparation; the novice will assume that what he sees in the microscope are real features of the original specimen.

As an example, a knife-cut section should be strain-free and free from marks resulting from imperfections of the knife edge (longitudinal features) or judder resulting from vibration of the knife edge producing marking transverse to the cutting direction. Excessive residual strain renders a section unsuitable for polarised light work or interference microscopy and creates difficulty in handling during mounting. Edge imperfections and judder introduce spurious 'features' to the cut section. Figure 1.1 shows two consecutive sections of part of a polypropylene moulding, one more carefully cut than the other. The poor section illustrates all the defects mentioned above and is virtually useless for subsequent critical examination.

2 A. D. Curson

(b)

FIG. 1.1. (a) Poor and (b) good thin sections of polypropylene.

The range of specimen preparation equipment available must be suited to the range of materials that are to be examined. Some materials respond quite successfully to traditional microtomy but there is the need to decide on the range of knives to be made available (e.g. glass, steel, tungsten carbide tipped). Selection of the profile of the knife is also important: plano-concave, plano-plano, chisel edge, wide angle, narrow angle etc. - they all have their individual applications, advantages and disadvantages. Other materials will require preparation methods involving embedding, lapping and polishing.

Should the general requirements of a laboratory call for lapping and polishing or any other powder work, it is advisable to separate these from all other operations. It requires only one particle of abrasive on a lens inadvertently wiped with a lens tissue to end the useful life of that lens. Microtomes operate on slides machined to tolerances that allow one to cut slices as thin as 0·2 .urn. If the slides are contaminated with abrasive powder particles this capability will not be maintained for long. Contamination of the mounted sections will also lead to incorrect interpretation and identification of possible additives and contamin-

Specimen Preparation 3

ation in the original specimen, particularly when using dark field microscopy.

It is also well recognised, and worth remembering, that in polymer microscopy the ratio of the time required for specimen preparation to that needed for actual microscopy is between lO: 1 and 100: I depending on the type of material and the form of specimen being examined. For sensible microscopy there is no such thing as a 'quick' section: it is either 'good' or 'bad'.

In classifying the different specimen preparation techniques it is tempting to make the subdivision according to the types of material, but experience suggests that this concept is not useful. This stems partly from an inability to define unambiguously such terms as soft, hard, brittle and ductile, when dealing with the modem range of polymers, copolymers, blends and composites. Also it is unhelpful to talk about specific polymers especially where the work is being carried out in the realm of the plastics industry. What might be supplied commercially as, say, polypropylene or unplasticised polyvinylchloride is likely to have no more than about 70% actual polymer content, and on occasions as little as 30%, with the remainder made up of particulate filler, fibres, pigments and other additives incorporated for stabilisation, lubrication or nucleation. Besides which, to work on the principle of classification by materials would not only lead to a lot of repetition but would also give the erroneous impression that there are 'standard' techniques and methods to be used for each polymer.

The intention is to describe different methods of preparation, with examples of their application, and leave it to the microscopists to decide which suits their purpose at any given time.

1.2 INITIAL APPROACH TO PREPARING THE SPECIMEN

When faced with a demand or need for microscopical work there are a number of basic rules which should always be observed:

l. Handle the sample as little as necessary and use forceps where possible.

2. Define the problem and identify which part of the sample requires e'\amination.

3. Identify the problems associated with obtaining the specimen in the correct form.

4 A. D. Curson

4. Assess the size of possible structural features to be examined and the areas over which this size might reasonably be expected to vary. It may be necessary to cut a 'sighting' section as an aid to making this assessment.

Such an approach should lead to a clear indication of the type of specimen to be prepared. It should fall into one or more of the following categories:

- Area of specimen: (a) large (b) small

- Surface: (a) as received (b) deliberately fractured (c) lapped or polished (d) cut

- Section: (a) thin (less than 15.um) (b) thick

- Powder dispersion - Melt pressing - Solid-liquid interface

1.3 AREA OF THE SPECIMEN

The surface area of the specimen is determined by two factors, both of which are dictated by the statistics of the situation. The first is the size of the structural elements that make up the texture of the specimen. The specimen area needs to be large enough to accommodate sufficient numbers of these elements so that the microscopist may be reasonably certain that the image is truly representative. Secondly, the area must be large enough to cover the area of the sample over which significant fluctuations in texture are likely to occur.

As an example, when examining the gelation mechanism of PVC within an extruder it would be necessary initially to cut a full section of the extruder core material which may be many square centimetres in cross-sectional area. This would give information relating to the variation in powder granule compaction and distortion as a function of its position within the extruder. To examine the individual granules in


greater detail it would, however, be necessary to cut much smaller (and thinner) sections from specifically selected areas as determined from the larger section.

1.4 SURFACES

There are many types and origins of surfaces that may need to be examined, the most common being:

-Mouldings - Extrudates - Films - Replicas - Powders and fibres - Fractures

(a) in-service failure (b) induced

-Cut -Lapped - Polished

The first five of these are 'natural' surfaces, i.e. they are surfaces that are a consequence of the manufacture of the sample. Fractures may be either natural (in-service failure) or induced as part of the specimen preparation procedure. The last three (cut, lapped, polished) are specifically 'manufactured' for subsequent microscopy.

Natural surfaces should not be handled. Contamination on the surface due to handling will lead to difficulty in subsequent specimen preparation and possible confusion in the interpretation of the image seen in the microscope. Any attempt to use even the mildest of solvents to remove fingerprints and the like may also remove important information relating to the original surface texture. In a few instances, such as in-service fracture of land drainage pipe, where the fracture surface might be obscured by soil, careful washing with distilled water in an ultrasonic bath may be necessary but the use of hydrocarbon solvents should be avoided at all times.

The study of most natural surfaces should start with an examination of the virgin surface. Therefore it is initially necessary only to mount the sample in a suitable way for presentation to the microscope objective, but ready for possible further preparation. In this respect some thought

6 A. D. Curson

ought to be given to supplementary examination if the information from light microscopy needs to be augmented by other methods. For example, if particularly fine detail is suspected or if the surface is obviously rough, scanning electron microscopy may be necessary to complement the light microscopical examination. In this case the sample mounting and subsequent preparation should be compatible with both types of technique and instrumentation.

1.4.1 Mouldings, Extrudates and In-service Fractures In the examination of the surfaces of mouldings and extrudates the initial aim is to present to the microscope objective a flat horizontal surface. Obviously, if the surface is not flat then a thin film replica may have to be made as described later.

There are a number of methods of getting the surface oriented in the required way, the simplest being to place the selected piece of sample on a microscope slide (assuming that the surfaces are parallel). A quick check through the microscope will soon confirm whether this method is satisfactory. It if is, the sample may be either left loose on the slide surface or held in place with a piece of double-sided adhesive tape.

If it is found that some adjustment, or tilting, of the surface is necessary, it is advantageous to put a small lump of modelling clay between the slide and the sample. The effect of small deflections of various extremities of the sample can be monitored through the

FIG. 1.2. Double tilt device for levelling surfaces.


microscope until the surface is considered to be horizontal. However, this procedure can be very tedious and frustrating; it can be very much simplified by using a levelling press.

Concern over the possible consequences of touching a virgin surface can be catered for by using a double tilt device such as that shown in Fig. 1.2. This enables the sample to be clamped at its edge surfaces only and to be tilted about two axes at right angles to each other. The example shown was adapted from a microtome accessory.

1.4.2 Films With the exception of samples that are fragile, e.g. thin film replicas, and those that may stretch under small stresses, by far the best method of preparing a film surface is to mount it on a jig of the type shown in Fig. 1.3. The sequence of events is illustrated; if followed conscientiously it results in a flat surface ready for direct examination or metallising. Apart from the flatness of the surface presented to the microscope, the main advantage of this technique is the minimal amount of handling of the actual surface that is necessary. Whether or not the surface presented is parallel to the focal plane of the microscope objective depends upon the accuracy to which the jig can be made.

Films such as replicas, which may not withstand this type of mounting, need to be attached to a microscope slide. This can be done by fixing the film in position using small pieces of adhesive tape,

FIG. 1.3. Mounting a thin film prior to direct examination or metallising.

8 A. D. Curson

double-sided adhesive tape or a suitable non-reactive adhesive such as natural Canada balsam, i.e. without added xylene. Mounting procedures are illustrated in Fig. 1.4. The major disadvantage of these methods is the difficulty in getting the film specimen to lie flat on the surface of the slide. Dust particles, irregularities in the lower surface of the film, air trapped between the sample and the adhesive, natural curling of the film, all contribute to produce a surface that is other than flat. In very difficult cases excessive handling of the specimen is required which could lead to damage or contamination of the surface to be examined.

If a liquid adhesive is to be used, metallising of the film surface should be carried out before sticking it to the microscope slide. This can be done by aluminising a piece of film many times larger than the selected area. The selected area can then be cut out using a scalpel or a razor blade and mounted on the microscope slide.

Replication of Surfaces Some surfaces, because of their curvature or inaccessibility, cannot be examined directly under the microscope. In such cases a replica has to be made which is subsequently treated either as a moulding surface or as a film surface. Also, to obtain the highest lateral resolution of surface structure it is necessary to take a thin film replica of the surface and to

FIG. 1.4. Two ways of mounting a film sample on a glass slide: (a) using strips of adhesive tape; (b, c) using a liquid adhesive.


treat this, after 'shadowing' (see later), as a thin section for examination by transmitted light microscopy.

Among the most common replication materials in use are:

1. Acrylic cements (polymethylmethacrylate) 2. Solution (1 %) of polystyrene in toluene or benzene 3. Solution of polyvinyl formal in chloroform or dichloroethane 4. Solution of polyvinyl alcohol (PVA) in distilled water 5. Low molecular weight acrylic sheet

The first three in this list involve hydrocarbon solvents which at best may simply remove features from the surfaces by solvation (e.g. removal of migrated additives) and at worst may alter the surface topography of the polymer itselfby solvent action. It is therefore advisable to limit their use to replication of the surfaces of processing equipment (e.g. the surfaces of casting rollers).

Water soluble PYA has the advantage that such active solvents are not present; nevertheless, some additives that can be present on the surfaces of plastic articles are water soluble and their possible removal should be noted.

The use of 'soft' (low molecular weight) acrylic sheet overcomes the problems associated with solvent action but it requires the surface that is being replicated to withstand temperatures of up to 70° C under moderate pressure.

Application of Replicating Solutions Whichever compound is chosen, the consistency of the solution must be that of a low viscosity syrup. It must wet the surface, spread or be spread easily and uniformly, and contain no bubbles. The viscosity can be adjusted by adding more solvent or solute as required. With proprietary brands of acrylic cements the viscosity is pre-set by the recommended mixing proportions of the constituents. In such cases the manufacturer should be consulted as to which grades are most suitable.

Invariably, air will be trapped by the stirring action during the mixing of the solutions. This can be removed by evacuation via a rotary vane vacuum pump. The roughing line on such a set-up should be interrupted by a cold vapour trap to avoid contaminating the rotary pump oil.

Application is preferably by using the natural wetting of the surface by the solution and aiding it by tilting the surface. If the surface being replicated is fixed and possibly at some angle other than horizontal then a soft-haired paint brush or a bar fashioned out of a small diameter

10 A. D. Curson

(1-2 mm) glass rod or tube may be used to assist spreading, as illustrated in Fig. 1.5. Whichever method is used, care must be taken not to disturb the sample/solution interface and to get a uniform coverage of the area of interest.

The production of a thin film replica is to be preferred to a solid block. Thin films have certain advantages:

1. They are quicker to dry from solution. 2. In reactive compounds they are much less likely to produce

internal bubbling as a result of exothermic reactions. 3. They are more easily removed from the surface. 4. Thin replicas of curved or convoluted surfaces can be laid flat to

facilitate subsequent microscopy and photomicrography.

For unknown surfaces it is worth experimenting on an area away from that of interest to ensure that the replica can be easily stripped from the surface. If too much force is required to peel the replica then it may be damaged, resulting in misleading information. When the replica is dry it can be gently stripped from the original surface and treated either as a film surface or as a thin section.

The most common fault in the preparation of thin film replicas is overestimation of the thickness of the film required.

FIG. 1.5. Spreading the replicating solution using a glass rod.


Use of Soft Acrylic Sheet This method of replication is to place in an oven at 60-70° C a piece of low molecular weight acrylic sheet, 2-3 mm thick and just a little larger than the area to be replicated, together with the sample. Mter about 10 min the sample is placed face downwards on the acrylic sheet and a suitable weight is put on top of it. After a further 5 min the oven is switched off and the contents are allowed to cool to room temperature. The weight and sample are removed and a replica of the sample surface will be found on the acrylic sheet.

1.4.3 Vacuum Metallising Only a few polymers lend themselves to direct high resolution reflected light microscopy. The more satisfactory materials (e.g. polymethylmethacrylate, polystyrene) can usefully be examined using bright field, dark ground and interference systems. Generally, though, the deposition of aluminium or silver on the surface at normal incidence increases the reflectance of the surface and improves the image contrast.

For a general improvement in image contrast, metallising to give 70-90% reflectivity is sufficient, the actual value not being critical. However, if a multi-beam interferometer objective is being used (see Chapter 6), care must be taken to try to match, as closely as possible, the reflectance of the sample with that of the reference mirror, or with one of the range of reference 'flats' available. In normal incidence metallising by evaporation it is good practice to use an extended source by employing a triple loop tungsten wire filament and subdividing the aluminium or silver wire between the loops. This ensures reasonable uniform coverage of the surface regardless of its topography.

Mounting of the metallised sample is exactly the same as for unmetallised samples.

Should it be necessary subsequently to remove an aluminium coating from the surface of a sample, this can be done by immersing the sample in an aqueous solution of ferric chloride. This technique is also useful for exposing the polymer film surface below commercially applied coatings.

1.4.4 S~adow Metallising Measurements of the height of discrete entities on a surface can be made by metallising the film or replica at(i known angle. In this case a single V-notch filament is used to provide a source approximating to a point source. The sample is positioned in the vacuum chamber to give the

12 A. D. Curson

selected angle of shadowing, the value of which depends on the roughness of the surface being studied but is usually 14°, giving a shadow/structure height ratio of 4: 1; see Fig. 1.6.

The shadowed replica or film is placed between a standard microscope slide and coverslip and immersed in a liquid matching the refractive index of the polymer or replicating medium. The preparation can now be examined using transmitted light microscopy.

1.4.5 Powders and Fibres The surfaces of powder particles and fibres are generally best studied using scanning electron microscopy. This is because the detail on such surfaces normally demands high resolution microscopy. As the resolving power of a light microscope increases, the depth of field decreases to the extent that for such subjects as powders and fibres only a very small part of the surface is in focus at anyone position of the objective or stage. Scanning electron microscopy has the advantage of a much greater depth of field.

There may be occasions when low resolution is adequate. In such cases a dry dispersion of the powder or of short lengths of the fibre on the surface of a microscope slide is all that is necessary. Any problems usually arise from ensuring that the range of particles seen in the field of view is representative of the sample as a whole.

Ignoring the problem of how representative a 20 g sample is of the contents of a 10 tonne silo, the following comments are confined to

Vacuum Chamber Film Deposited Aluminium

Deposited Aluminium

x/ y = 4/. Slide Immersion Liquid

FIG. l.6. Shadow metallising and subsequent mounting of film samples.


ensuring that what appears on the microscope slide is representative of the 20 g sample. The temptation to use sprays, air agitation, dispersions in liquids, and the like should be resisted. They are very prone to problems of segregation leading to significant sampling bias.

The simplest and most effective method of preparing a powder sample is illustrated in Fig. 1.7 where the sample is shown being first gently stirred with a spatula. The sample container must not be rolled or shaken since this can result in size segregation, the smaller particles accumulating at the bottom of the container. A small sample is removed from the container and placed on a clean microscope slide. This pile is then spread out using the point of a needle, and finally a uniform distribution is achieved by gently tapping the underside of the glass slide. Short lengths of fibres can be treated in much the same way or, alternatively, several lengths or a monolayer bundle of the fibres can be fixed with adhesive tape at either end to the microscope slide.

A final tip on the preparation of powder and fibre samples for surface examination can be borrowed from the textile industry. Unless there is a need to examine the silhouette of the individual entities it is advisable to place the prepared slide on a backing having colouring similar to that of the sample itself. This will reduce the glare from the background and enable better appreciation of the variations of contrast on the surface of the sample.

FIG. 1.7. Preparing a dispersion of a small sample of powder.

14 A. D. Curson

1.4.6 Induced or Controlled Fractures Although cold fracturing is generally described as an electron microscopy preparation technique, this should not be thought of as a reason for excluding it from light microscopy practice. Indeed with some modem composites and filled materials it can be the only method of revealing the bulk structure of the sample. It has been found to be useful in revealing subsurface voiding in mineral filled nylon mouldings. Traditional microtomy could not be used because of the hardness of the filler, and polishing techniques modified the prepared surface sufficiently to destroy or conceal evidence of the presence of the voids.

There are no hard and fast rules governing when this technique should be used, except when it is suspected that other preparation techniques are not providing a sufficiently complete picture of the internal texture. However, it does have three big disadvantages. First, it does not necessarily produce a flat smooth surface, making high resolution light microscopy difficult. Secondly, there is the danger of misinterpreting fracture morphology as internal texture. Thirdly, as a fracture path it will follow the line of least resistance. The texture revealed may not, therefore, be representative of the bulk structure.

The method is to make a saw cut about I mm or 2 mm deep on the edge of the sample and at one end of the line of the intended fracture. The sample is then immersed in liquid nitrogen for at least I hour for samples up to 5 mm thick, longer for bulky specimens. Shorter times will result in ductile fracture of the sample, particularly in the central regions, the morphology of which will totally destroy the original bulk texture of the material. The sample is then removed from the liquid nitrogen and placed on a cold, hard, firm surface. The blade of a cold chisel is put into the saw cut and given a sharp tap with a hammer.

The two pieces must now be allowed to come back to room temperature and left to dry. They may then be treated in the same way as other surfaces and examined directly, metallised or replicated.

In the examination of all fracture surfaces, whether natural or induced as part of the preparation technique, it is important to examine both halves ofthe fracture. Failure to do so can yield misleading results. For example, a cavity on one half may be represented by a cavity on the other surface, in which case it may reasonably be assumed to be a void. Such a cavity may, however, have a protruding particle at its equivalent position on the second surface, giving rise to a totally different interpretation.


1.4.7 Cut Surfaces This technique has very limited application. The areas in which it will be found most useful is the study of foam structure and porosity in large samples (see Section 1.7). In some instances it may be considered necessary to prepare cut surfaces of solid materials, e.g. if etching programmes are to be pursued. These would be prepared as for thin sectioning of the sample, with the block held in the microtome vice being retained for examination.

For the study of foam structure, a steel straight edge and either a fresh single-edged blade or a scalpel is required. With the steel edge as guide, and using the point of the blade, consecutive light cuts are made, progressing deeper and deeper into the sample until the two parts are severed. To try to force the blade through too quickly will distort the structure of the specimen and produce a non-flat surface.

1.4.8 Lapped and Polished Surfaces Modern materials, especially composites, call for preparation techniques in a class of their own. The hardness of the fillers and fibres used make traditional polymer preparation methods quite inapplicable, and the techniques used in petrology and mineralogy have had to be 'poached' and modified to fill the requirements. Such methods will be found useful in the assessment of impregnation of polymeric and nonpolymeric matrices between fibres, the assessment of internal damage in test specimens, the assessment of uniformity of fibre distribution, and the monitoring and measurement of lay-up angles in multilayer composites.

Polishing is an extension of the lapping process and should be restricted to materials in which both matrix and filler are hard, and to those situations where it is essential to provide fine detail information. The reason for this is that the polishing stage is lengthy and, unless it is the only way of revealing the detail required, would add unnecessarily to the preparation time.

The sample (or samples) can be worked dry or embedded. The advantage of embedding is that it reduces rounding-off of the edges of the specimens, although this can also be avoided by sandwiching several dry samples together between two waste cheek plates. Embedding will be essential if the samples are thin and therefore in need of some support. Non-embedment, or the use of a suitable resin which can be subsequently removed (e.g. acrylic later removed with chloroform), is to

16 A. D. Curson

be preferred in the preparation of specimens in which the degree of internal damage is to be assessed. If a non-extractable embedding resin is used in such circumstances there is the danger that it will infill some of the damage structures and thereby reduce their contrast in the final microscopic image. However, given that the necessary microscopic techniques (e.g. interference contrast) are readily available, such infilling can be identified and its presence used, with caution, to distinguish those features that were open to the original surfaces of the sample from those that were totally enclosed. Suitable resins are any low viscosity cold-setting epoxy types. Some acrylic compounds are also suitable but not those with a powder component.

For all preliminary cutting and shaping it is necessary to have a diamond-edged saw available. Hacksaws and the like are second best; not only do they become blunt quickly but they cause severe damage to the specimen extending well beyond the depth to which subsequent lapping and polishing will proceed.

Both lapping and polishing are carried out either manually or with machines built for the purpose. However, there is little if any preparation equipment designed specifically for such work with polymers and their composites, so it is necessary to proceed with care and to remember that the techniques being used were probably designed to cope with rock and metallic specimens.

For some critical applications where flatness of the surface is essential hand polishing using soft cloths should be avoided as this inevitably gives rise to 'profiling' (uneven wear of different constituents and at open phase and grain boundaries).

The Manual System The inclined bench is loaded with four grades of 'wet-and-dry' silicon carbide paper and a weir of water is produced to flood each one, keeping it wet and removing debris. Suitable grades are 220, 320, 500 and 1000. The coarsest grade, 220, is used initially to expose the full surface of the specimen since it is likely that there will be some embedding resin to be removed from the surface.

This technique requires firm but not heavy pressure on the specimen which is drawn down the slope of the bench towards the operator. The specimen should be returned to the top of the plane without touching the paper. Between consecutive 'working' strokes the sample should be rotated through about 45° to ensure uniform abrasion of the whole surface. A scrubbing action must be avoided as this will lead to a nonflat finish. Working towards the finest grade, use each stage only


sufficiently to remove the abrasion marks due to the previous stage. On the final grade of paper (1000) the pressure on the sample should be gradually decreased until the last few strokes are almost aquaplaning over the paper. A 'lapped' surface will now have been produced which, after drying, will be suitable for examination for fairly large detail such as interlaminar splitting or large cavities.

Mter thorough washing the specimen can now be polished. This is done using a high quality nap finished cloth laid on a glass plate. The cloth is thoroughly soaked with water and aluminium oxide (0·3 /.lm grade) is sprinkled on the surface. The specimen is placed face down on the cloth and a circular motion maintained, moving the specimen around so as to use the whole of the surface of the cloth. The specimen should also be rotated during this action. The pressure on the specimen should not be excessive and the cloth should be kept wet. It is inadvisable to use too much polishing powder since an excess will tend to impair the finish of the final polish. The sample should be washed, dried and inspected regularly until a satisfactory finish is obtained.

The Motor Driven Systems Generally speaking, for all but the hardest materials, machines that utilise a slurry of abrasive powder and liquid carrier, e.g. silicon carbide and water, are unsuitable for polymer based samples. This is because individual particles of the abrasive can become embedded in the relatively soft plastic component or wedged in the filler/matrix interface. They can be dislodged during a subsequent stage, thereby contaminating the finer abrasive, or they can remain in position and obscure the final microscopic detail of the surface. In this latter respect a few particles can be tolerated but it is not unusual to find more than 60% of the surface contaminated by such means.

Carbon fibre filled polyetheretherketone reacts favourably to the following sequence:

(1) Initial 'dressing' (flattening) of the surface using the 220 strip on the manual system

(2) 5 min using 600 grit silicon carbide in water on a cast iron plate (3) 5 min using 9 /.lm aluminium oxide in water on a cast iron plate (4) 5 min using 3· 5 /.lm aluminium oxide in water on a cast iron plate (5) 30 min using 6·5 /.lm diamond paste with recommended carrier

on soft solderllead plate (6) 30 min using I /.lm diamond paste with recommended carrier on

soft solderllead plate

18 A. D. Curson

The timing indicated is not necessarily reproducible and will vary from sample to sample depending on the properties of the polymer system being worked (including direction of fibre orientation), the total area being polished and the experience of the operator.

Each stage is used until the abrasion marks due to the previous stage have beenjust removed. The sample, the sample carrier, the plate and the slurry feed mechanism must be thoroughly cleaned between stages during the lapping process on the cast iron plate. In the polishing process a separate soft solder or lead plate is provided for each grade of diamond, so it is only the sample(s) and carrier that need to be cleaned.

Using the sequence described and the equipment shown, surfaces having a high polish and with a high degree of flatness with no discernible profiling have been produced.

With samples in which impregnation by the abrasive is a problem the surface of the cast iron plate can be covered with a self-adhesive fixed abrasive paper and water is fed continuously on to its surface. The paper is changed for a finer grade when necessary, with little more than general washing required for the sample. However, this fixed abrasive lapping does not produce a surface flat enough to be adequately polished on soft metal plates; a more amenable system has to be used, such as colloidal suspensions of polishing media on pressed unwoven cloths. With this technique there is some danger of profiling but not as great as that experienced with the manual system.

With highly polished surfaces there is normally no need to metallise or otherwise further prepare the surface. The variation in reflectivity between the components of the sample is sufficient, even in polymer blends, to enable them to be identified, particularly when using reflected polarized light microscopy.

1.5 SECTIONING (SECTIONAL SLICES)

The purpose of taking sections is to examine the internal or bulk structure of the sample. The thickness of the section, whatever its area (see Section 1.3), is determined by the size of the structures to be examined and, to a lesser extent, by their concentration. A good working rule is to divide the size of the structures by 4 and cut sections of this thickness. However, this can present problems, one of which occurs when the calculation calls for a section thickness ofless than I J.lm and


another when the structure varies considerably within the sample. In the first case it is by no means impossible to section as thin as

0.5 11m or less, but to do so requires not only expensive instrumentation but considerable operator skill and experience. Where either or both of these resources are absent the answer is to section 'as thin as possible' and hope that 'optical sectioning' with the microscope will achieve the desired result, but too much reliance must not be placed on this approach as it has severe limitations and can lead to slipshod habits resulting in poor microscopy.

To section too thin with respect to structure size will often result in lack of contrast and detail in the image. Therefore, for samples containing a wide range of structure sizes it is good practice to section the whole area according to the thickness criterion determined by the largest structure. This can then be used to locate the positions of the finer texture which are identified on the original sample and individually sectioned at the reduced thickness. It is poor practice to study, in detail, both coarse and fine structure in the same section.

In a few instances an overriding factor governing the thickness of the section to be cut will be the birefringence of the crystalline structure being examined. For some aspects of polarized light microscopy, e.g. depolarized light intensity analysis of nylon, it is advisable to have the optical path difference generated by the sample less than one order. Since this path difference is the product of the birefringence and the specimen thickness, and the birefringence is constant, the only controlling variable is the section thickness.

In many cases, if not all, it is worthwhile cutting at least two, preferably consecutive, sections and mounting one in a medium matching the refractive index of the polymeric matrix and t~e other in a medium of known mismatch. This makes the task of identifying voids and carrying out refractive index measurements of the sample material much easier.

1.5.1 Microtomes For polymer work the best and most reliable microtome, in terms of consistently good results, is the base sledge type. Sections will be cut as thin as 0.5 11m, so rigidity of all parts is essential. This applies not only to the runners, or glides, on which the vice rides but also to the knife blade itself.

Building the sledge in the vertical direction and operating the vice movement via a rotating handle is an alternative design and one that the

20 A. D. Curson

author finds preferable since the control of the movement of the sample seems more positive. Nevertheless the standard horizontal base sledge has probably the greater number of devotees. The main disadvantage of the rotary, vertical microtome is that it presents problems when attempting ice embedding, but these can be overcome.

Not all microtomes are suited for accepting tungsten carbide tipped knives. Many of the sturdier base sledge microtomes, which with their rotary counterparts are the only ones really suited for polymer work, will accept only knives of 170 mm or longer.

The basic design of most microtomes appears to be based on the requirements of the biologist. In particular, clamping arrangements are such that it can be impossible to section specific samples in the required direction, and modifications to the equipment may need to be made (e.g. the manufacture of special vices).

Cold (and Hot) Stage Microtomes Mention will be made of semiconductor devices used for embedding in ice. Another of their applications is in reducing the temperature of the sample sufficiently to make it less flexible and therefore easier to cut. A good rule-of-thumb is that the temperature at which the material is cut should be approximately 30° C below its glass transition temperature (Tg ).

Semiconductor stages reduce the temperature of a small sample to about - 30° C but there are occasions when it is necessary to cool the specimen (and the knife at such low temperatures) well below this. Sectioning of polytetrafluoroethylene (PTFE) and plasticised PVC are two examples. There are very few commercially available instruments that will cope conveniently with this situation but suitable systems based on cooling with liquid nitrogen can be built in-house. The instrumentation illustrated in Fig. 1.8 will take the sample and the knife down to at least -160° C.

On the other hand, it may be necessary to raise the temperature of a sample to soften it before successful sectioning is possible. This may be done conveniently by blowing hot air over the knife and specimen (a hair dryer is suitable). However, great care must be taken not to modify the polymer texture by overheating.

1.5.2 Microtome Knives The design of commercially available microtomes means that the microtomist is faced with forcing a stationary blade through a solid


FIG. 1.8. Low temperature microtome: (a, b) liquid nitrogen reservoirs; (c) base sledge microtome; (d) heater controls.

block of material. The fact that he is removing a very thin slice eases the problem~ Inclining the knife does little to help in all except two instances - generally it only enables one to damage a greater length of the knife edge on the same width of sample. It also tends to promote distortion and curling of the section. The two exceptions are the sectioning of circular profiles and where the length of cut is equal to or shorter than its width. In these cases less force is required to cut the section using an inclined knife edge.

The fundamental principle underlying microtome design is to ease the knife edge through the material to produce what is in reality a controlled fracture path. The smoothness and linearity of this 'cut' will depend upon the sharpness of the knife, its rigidity, the geometry of the knife edge with respect to the direction of propagation of the cut, the speed with which the cut is being made, and the rigidity of the sample.

Types of Knife There are three main types of knife used in polymer microtomy:

Steel. These are the traditional microtome knives. They are ideal for the soft plastics such as polyethylene and polypropylene as long as these are unfilled and unpigmented. They should also be used when sections wider than 3 mm are required. Their disadvantage is that they readily

22 A. D. Curson

blunt and require regular honing or stropping. In spite of this the edge soon becomes irregular, leading to unacceptable knife marking of the sections.

Tungsten carbide tipped (TeT). For wide st:ctions these are to be preferred to their steel equivalent as they retain their edge for a greater time. They can also be used on materials incorporating the softer fillers, but silica, glass fibre (and beads) and carbon fibre filled materials should still be avoided. Their disadvantage is their high price.

Glass. For fine work on unfilled materials these are ideal. Freshly broken glass offers the sharpest edge available, and on a 45° knife the rigidity is also present. The disadvantages are the difficulty in producing a straight edge that is perpendicular to the sides of the knife, and the extreme brittleness of the edge. As a result of the latter, the technique of using such knives is to dress the sample with one knife and to use a new knife to cut the actual section. This should be taken within the first five or six cuts; otherwise the knife should be changed again. This knife can then be used to dress the next sample, and so on. Experience has shown that glass plate of between 4 mm and 6 mm is most suited to this application, thereby restricting the area of the cut section to a few square millimetres (the smaller the better). Proprietary glass knife clamps are found wanting in this application; a more suitable clamp is shown in Fig. 1.9.

FIG. 1.9. Glass knife holder that can utilise the movability of the steel knife holders.


Sharpness of Knife Edge The sharpness of an edge is defined by its radius of CUIVature and the uniformity of this radius along the length of the knife edge. Localised variations in the CUIVature produce unacceptable knife marks such as those illustrated in Fig. 1.1 (a). Specialised equipment and experience are required to keep the edge of a steel (or tungsten carbide tipped) knife in a suitable condition. Invariably this entails returning it to the manufacturer. Few, if any, proprietary knife sharpening machines readily produce an edge of acceptable quality for polymer sectioning, although, given sufficient time, a dedicated operator may eventually develop the necessary skill and technique.

Rigidity of the Knife If the knife blade is too flexible it will distort under the pressure of cutting, resulting in an uneven thickness of cut, possibly with eventual riding up to the top of the sample. It may even cause permanent distortion of the knife edge. For this reason, when selecting steel or TCT knives, the choice is restricted to those with either a plano-plano or a chisel-edge profile (see Fig. 1.10).

Geometry of the Knife Edge The lowest face of the knife blade should be raised only a few degrees (5° at the most) to the plane of the intended cut. An excessive angle will result in knife 'judder' producing marks similar to those indicated in Fig. 1.1 (a). The severity of such marks can in some cases be sufficient to obscure structure.

Setting the knife at the correct angle is straightforward when using glass knives but is more problematical when employing steel or TCT

Plano - Concave (a,b) Chisel-Edge (d)

Plano· Plano (c) Glass

FIG. 1.10. Profiles of microtome knives.

24 A. D. Curson

Incorrect Setting

Sample Surface

Correct Setting

FIG. 1.11. Setting the knife angle for steel and TCT knives.

knives. This arises from the need to take account of the small honing bevels to be found on such knives and which are not easily discernible to the naked eye. The problem and its solution is illustrated in Fig. 1.11. The solution is to start with an angle of about 5° between the back of the knife and the surface of the sample and to attempt to take consecutive sections of 1 pm thickness. The angle between the knife and the sample should then be increased by no more than 1 ° or 2° increments until a section is produced with each pass of the sample. The smallest angle which the knife makes with the surface of the specimen making this possible is the correct angle for that knife.

1.5.3 Speed of the Cut There are no strict guidelines for prejudging the correct speed at which a cut should be made. It will vary from sample to sample and will have some dependence on the geometry of the knife edge. When experiencing difficulty in producing a good section it is worthwhile experimenting with different cutting speeds.


1.5.4 Rigidity of the Sample By whatever means the sample is prepared for holding in the microtome (see later) it is important to ensure that only a small amount projects above the clamping surface. If too much of the specimen is not given adequate support, not only will it be difficult to cut sections of consistent thickness but any of a number of other defects will be built into the section. A good working rule is that the height of the material (specimen plus clamping aids) projecting above the clamping surface should be about two-thirds of its width between the clamp jaws.

1.5.5 Thin Sectioning: Holding the Sample Having decided on the thickness and area of the section to be cut, the next problem is to find a suitable way of holding the specimen in the microtome. It is best to consider this according to the form of the specimen:

- Powders - Fibres - Granules - Mouldings and extrudates -Foams - Films - Composites and 'hard' materials

Obviously there will be occasions when a particular sample cannot be identified with any of these classifications or it ought to be considered in a different class from that which at first sight is apparent.

Powders The only satisfactory way to section powder particles is to embed them. Three main types of embedding material can be considered.

Epoxy resins. These need to be of very low viscosity with good wetting characteristics, ensuring a high level of penetration into very small pores. There are a number of suitable resins available and, apart from the properties mentioned above, it is worthwhile checking on the shelf life both before and after the opening containers, the reliability and tolerance to variations in the recommended proportions, and also the effect of absorbing water from the atmosphere. The advantages of epoxy resins are that they provide good penetration, especially where friable and fragile powders are involved, and they are less likely to be attacked by mounting liquids.

26 A. D. Curson

Acrylic cements. The use of systems based on acrylic powder plus monomer is not recommended for powder embedding since the structure of the acrylic powder particles in these materials is not lost. The best acrylic systems are the low viscosity two-part liquid mixes; they have quite good penetration which can be aided by vacuum cycling. The main advantage of embedding in acrylic is that, should it be necessary to remove the embedding material from the particle section, this is easily done by using chloroform or acetone, accepting that neither solvent will attack the sample itself.

Air can be removed from epoxy or acrylic resin by vacuum cycling at least four times. The procedure is to put a small amount of the powder sample into a polyethylene or gelatine capsule and add a few drops of the resin, mix the two together (vacuum cycle the mixture if necessary) to ensure good penetration, and top the capsule with resin. Mter curing, the polyethylene capsule can be removed, but if gelatine has been used it will be permanently attached to the resin. The embedded powder can then be held in a suitable clamp on the microtome.

Ice. The use of a cold stage in the microtome vice is invaluable if sections need to be cut quickly, but subsequent handling is more difficult and there is the danger of damage to the internal structure of the particles owing to inadequate penetration by the water or expansion of the water on freezing. A number of semiconductor devices are available which will achieve a platform temperature of -20° C to - 30° C. With these devices it is a good idea to have the DC supply controlled by a water switch so that the power is automatically shut off if the flow of water stops.

The technique for using this equipment is to build up a platform of ice on a single layer of paper tissue placed on top of the cold surface, add the powder particles on top of this layer and encapsulate them in more ice using water added via a micro-pipette. The tissue paper provides a stronger bond to the semiconductor surface than would be obtained with ice alone. Sectioning is carried out in the usual way, generally using a glass knife. The use of a stereo microscope mounted above the microtome is advantageous since, as soon as the sections are cut, the ice melts and the sections are left free floating on the upper surface of the knife. It is possible also to freeze the knife but for this particular application this is not essential. The sections need to be allowed to dry before being mounted.

One must ensure, having finished using a cold stage on a microtome, that the equipment is thoroughly dried and oiled to prevent the formation of rust.


Fibres Fibres can be treated in much the same way as powders, with the fibres supported parallel to the longitudinal axis of the moulding capsule. On occasions it may be necessary to section fibres parallel to their long axis; this can be done by embedding the fibres in a sheet of acrylic. This is perhaps the only application for powder plus liquid acrylic systems which have the advantage that their viscosity increases rapidly very soon after mixing. Before it gets too viscous, two glass plates should be lightly smeared with petroleum jelly and a layer of the resin spread on the greased side of one. To this is added a layer of fibres aligned parallel to each ot1er, then some more resin, and finally the second plate with its greased si e downwards. Light pressure is applied to obtain a total resin thickness f 1-3 mm. When the resin has cured, the plates can be stripped fr~~ the acrylic which is then sectioned in a microtome with the fibres lfing in the required direction.

Granules \ As for powd~rs, encapsulation of granules in resin or ice is possible, the latter beinglpreferred. In the absence of freezing equipment, good results can br obtained by sticking the granule to the end of a short length of met~l rod using a low melting (about 60°C) dental wax. The rod is held in\the microtome by means of the capsule holder.

\ \

Mouldings and 1ixtrudates These are treate~ in identical ways and are normally held directly in the vice of the microtome. Some instruments have very coarse serrations on the jaws of their clamps and in such cases it is advisable to sandwich the sample, particularly if it is relatively thin, between two cheek pieces of scrap rigid polypropylene. If the sample is not flat and parallel sided then cheek pieces with suitable stopping steps are required to hold the specimen firmly.

Foams These are probably the most difficult materials to prepare. Ifthe cellular structure is open, or interconnected, embedding in epoxy will help. If the cells are closed, or unconnected, the problem is magnified ten-fold. The greatest difficulty is in clamping the sample in the microtome without damaging it. For this reason, and because many foam materials are soft, ice embedding is recommended. Very low temperatures may be necessary.

28 A. D. Curson

Films The thinness of such samples should not deter the microscopist from attempting to section them. The secret of success is to mount the film in such a way that what is presented on the microtome knife is a relatively thick, solid specimen. This can be done by embedding a piece ofthe film on edge in ice, or in an epoxy or acrylic or any other suitable resin. A more expedient solution is to sandwich the film between two cheek pieces of 3 mm thick rigid polypropylene.

1.5.6 Some Hints on Thin Sectioning Technique The room in which thin sectioning is carried out should ideally be dustfree. Contamination of the sections will not only lead to confusion in the eventual interpretation of the microscopic image but may also prevent the coverslip from lying as flat as possible. This is a common cause of non-uniform focus both in the microscope image and in subsequent photomicrographs.

Slides and coverslips should be thoroughly cleaned and, in the case of coverslips, both slides should be scraped with the edge of a new razor blade. This latter action is essential as it removes the small pieces of glass found adhering to the surfaces when the coverslips are unpacked. Failure to remove them will result in the coverslip not lying flat.

When 'dressing' the sample the feed mechanism should not be advanced by more than 10 or 15 f..lm at a time, particularly when sectioning ice-embedded materials. The greater forces involved in cutting thicker slices may distort the sample or even dislodge it from its ice bed. The dressing stage should always be finished with several cuts of the thickness of the section to be taken. If the intention is to section a natural surface layer in the plane of the surface, 'dressing' is not relevant; the main concern is to get the surface and knife edge parallel before attempting to take the section.

If there is some natural direction associated with the sample, e.g. an extrusion or machine direction, or one related to injection moulding or other flow profile, the sample should be rotated so that the knife is cutting at a small angle to this direction. This enables structure relating to the processing conditions to be distinguished from any arising from sectioning faults.

The use of a fine, soft-haired brush or fine pointed forceps is invaluable in removing the section from the knife edge (see Fig. 1.12); it also helps to prevent curling of the section. However, extreme care should be taken when using either aid. Pulling with forceps tends to


FIG. 1.12. Removing a thin section with the help of a soft-haired brush.

damage and stretch the section, and heavy handedness with even a soft brush on polyethylene will leave severe brush marks on the section.

Applying a small amount of the mounting liquid, water or alcohol via a pipette, spray or bath to the knife edge during cutting will reduce 'judder' and make retrieval of the section easier.

The cut section of a moulding may be shorter than the length of the sample from which it has been removed. This may be due to compression by the cutting action: equally, it may be due to the relaxation of frozen-in processing stresses as the support of the bulk sample is removed from the section.

Unless it is absolutely necessary to retain the leading and trailing edges of the section, they should be trimmed offwith a new razor blade. These are the two most likely parts of the section where scrolling and fold-back will occur; incorporating them in the mounted section will prevent a flat preparation from being obtained.

Curling of the section is probably the most difficult to overcome. It arises from the differential relaxation of stress within the section. In polymers the various stress levels will be introduced either by the action of the microtome knife or, more commonly, by the processing conditions under which the sample was produced. Consequently no amount of soaking (or floating) of the section in warm oil or water will improve

30 A. D. Curson

matters. In fact, such action may only make the situation worse by encouraging even tighter curling. Curling due to poor microtomy technique can be minimised, if not eliminated, by careful attention to all the various points raised in this chapter. Curling due to the relaxation of frozen-in stresses can be reduced by cutting thinner sections.

The best solution to curling problems requires a degree of micromanipulation using a stereo microscope. The curled or corregated section is held reasonably flat under the microscope and a suitable slice is removed from across its width using a scalpel or razor blade. This is then cut into as many smaller pieces as are necessary to ensure that each will lie flat. In practice, for a moulding of wall thickness 3-4 mm this usually means three or, at most, five separate pieces each usually relating to specific zones of structure arising from the processing conditions. Each piece is mounted individually; not under the same coverslip.

To remove acrylic cement from very small thin sections, dry-mount the embedded section between a slide and coverslip. Keeping the slide horizontal, introduce the chloroform or acetone at one side of the coverslip using a fine pipette. When the liquid reaches the other side of the coverslip maintain a constant flow for a few seconds by soaking the solvent into the edge of a tissue. Stop the solvent feed and soak up as much of the remaining fluid as possible without disturbing the coverslip. Let the preparation dry out, and finally extract and mount the acrylic-free section using a stereo microscope.

When mounting thin sections, a single drop of mountant should be placed in the centre of the slide and a similar one on one face of the coverslip. The section is then placed on the drop on the slide, and the coverslip, drop downwards, is lowered gently on to it.

It is always good practice to minimise the amount of mountant required. It is the surface tension pulling the coverslip down on to the section that ensures a good, flat preparation. If there is too much liquid present the section will remain suspended and not flattened. It is much easier, and neater, to introduce a little more mountant using a fine pair offorceps loaded with liquid in much the same way as a draughtsman's pen than to attempt the more unsatisfactory operation of removing excess liquid using the edge of a tissue.

Ifhigh resolution microscopy is envisaged, or if the mounted section is to be stored, the coverslip should be 'ringed' or fixed with a suitable fixative.


1.5.7 Large Area Thick Sections There are a number of circumstances in which the demand is for low resolution microscopy but covering a large area. Two such instances would be the study of gelation processes of PVC within extruders and the glass fibre orientation in moulded rail clips. Because of the detailed information required, direct examination of a lapped surface is unsuitable and the softness of the polymer matrix precludes polishing. Thick sectioning, 20-30 Jlm for glass fibre filled materials and 40-70 Jlm for unplasticised PVC, provides ideal specimens for this type of work.

The technique is to produce a finely lapped finish on one surface of the sample, as described earlier, finishing on 1000 grade paper. This face is then cemented to the surface of a glass plate which may eventually be used as the microscope slide of the preparation and therefore needs to be of a suitable thickness (0·6-1·2 mm). The bonding material can be epoxy resin, in which case the glass surface needs only to be thoroughly cleaned, or it can be an acrylic cement, in which case the surface of the glass has to 'frosted' by working it on a suitable lapping machine. This frosting ensures adequate keying of the acrylic to the glass surface and prevents splitting of the interface at a later stage. A frosted surface can be used with the epoxy cement but this is inadvisable as it is likely to contribute confusing detail to the image.

When the adhesive has set, the excess bulk of the sample can be removed with a combination of saw and high speed router. With the latter (as with a circular saw) the supporting glass plate is held in position using a vacuum table or double-sided adhesive tape. Sensible use of such a high speed cutter (removing only very thin layers at each pass) does not produce significant alterations to polymer texture or fibre orientation. Spindle moulders, lathes and static cutter forming machines are less suitable since these do tend to produce excessive damage. Even with routers the design of the cutting edge of the tool appears to be important, pointed cutters being least satisfactory.

Although it is possible to complete the process with the router, the proximity of the high speed cutter to the glass surface suggests that it is prudent to stop this part of the operation leaving about 100 Jlm thickness of specimen on the plate. The remainder can be removed on the lapping bench or machine, monitoring the thickness with a micrometer and finishing with grade 1000 pa per. In the latter stages care must be taken to wear the section down evenly over its entire surface.

It is possible to carry out the entire thinning process on a lapping

32 A. D. Curson

machine if the necessary section preparation jig is available (such jigs ensure even lapping of the sample area). Such jigs do, however, restrict the area of specimen that can be prepared, although this is not insubstantial. The biggest problem is that with large areas there is insufficient load on the jig to ensure a reasonably fast rate of abrasion with polymeric specimens. Consequently the surface is subject to overheating and the time to complete the operation is unacceptably long. Furthermore, as the specimen gets thinner the lubricant (usually water) penetrates the glass/resin interface and severely affects the adhesion, often to the extent that the specimen works loose and is destroyed.

If any epoxy adhesive has been used, the specimen is completed by adding another suitable glass plate as the coverslip, using as mountant either the same resin or a suitable liquid matching the refractive index of the polymer. If an acrylic adhesive has been used this is removed by soaking in chloroform or acetone, whichever does not attack the specimen, and the clean section is mounted in a liquid with a matching refractive index.

The use of epoxy resin ensures that the specimen remains flat once the excess material has been removed. In some samples there will be considerable frozen-in stress which tends to distort the final section. This can be a problem if using the acrylic cement approach; the final mountant in this case needs to be natural Canada balsam and the mounted section must be kept under pressure until the balsam has hardened.

The disadvantages of the permanent epoxy system is that there is likely to be a fairly large refractive index mismatch at the interfaces; if these are rougher than expected a poor image will be obtained in the microscope. Also, with large areas it is not easy to ensure that the epoxy layer is free from bubbles arising from entrapped air or from incomplete infilling of cavities in the surface of the sample; these will detract from the ultimate quality of the preparation.

1.5.8 Small Area Thick Sections For small area samples the technique is modified by replacing the machining of the specimen with the use of a suitable jig on the lapping machine or by careful filing by hand. In the latter case the work is taken to the file (which is held horizontally in a vice) and not the file to the work.


1.5.9 Thin Sectioning of Composites and Brittle Materials To obtain satisfactory thin sections of such materials as carbon fibre filled polymers, both surfaces need to be polished and flat since their texture will contribute significantly to the microscopic image. The previous technique, for thick sections, is extended to produce a polished surface, but a very weak bond will be achieved if it is attempted to stick this face directly on to the surface of the glass plate. A more satisfactory approach is as follows.

Select a thin (0·8-1·0 mm) slide and put it on a hot-plate at about 60°C. Deposit on it four drops of a selected resin spaced so as to give eventual support to the sample (see Fig. 1.13). When cured (after 20-30 min), remove the slide and allow it to cool. Using a suitable device, e.g. a section preparation jig, grind the hardened pips down uniformly using 600 grit silicon carbide until their thickness together with that of the slide is no more than 1·2 mm. Clean and dry the slide and cement the polished surface of the specimen to the slide using the pips as supports (Fig. 1.14). Place a heavy weight on top or use spring loading to maintain positive contact with the tops of the pips, and cure at room temperature.

Proposed Resin Drops Position of Sample

Resin drops

\0 0 1 .... ""

'/ Slide

Plan View Side View

FIG. 1.13. Making the supporting pips for thin polished sections.

Heavy Weight Polished Surface

Slide

FIG. 1.14. Using the pips as supports prior to final thinning.

34 A. D. Curson

The remainder of the procedure is the same as that for large area thick sections but is extended to finish the top surface by polishing. Lapping should be taken only to a section thickness of no less than 25 J.lm, the final thinning being achieved by polishing. With care sections as thin as 7 J.lm of 30% carbon fibre filled polyetheretherketone have been produced by the author.

1.5.10 Sectioning Using Diamond Saws An alternative method of cutting sections as thin as 15 J.lm of brittle, hard or composite materials is to use an annular (or peripheral) diamond-eged saw. The specimen is mounted using a low melting point wax as illustrated in Figs 1.15 and 1.16 which show the use of an annular blade. The glass plate (G) is necessary to clean the blade edge of any firmly attached polymer debris; the double jet oflubricant (water) is essential to wash away loose debris.

Experimentation on waste pieces of material is required to establish the cutting speed that produces the best surface finish. This cutting speed will be a function of blade speed and pressure of the sample on the blade edge.

Using the correct speed, a surface is cut on the specimen. If thick sections are required, the sample (or blade) can be advanced the required distance (allowing for the thickness of the blade) and the section taken. Sections thinner than about 80 J.lm tend to curl and this becomes more severe with decreasing thickness. To overcome this problem, the first face of the section should be cut and then a glass

Saw Blade---------...

Glass IG)

Sample ........ "'"--L_ Dental Wax __ ~

5mm Glass Plate

Mounting Table

FIG. 1.15. Mounting a sample to be cut with an annular saw.

Specimen Preparation

saw8lad8J Lubricant

Double Jet

Top VIew of Lubricant Feed

~~~==::_ To Mains Water Supply

35

FIG. 1.16. Sample table and double lubricant jet in relation to the saw blade.

coverslip should be stuck to this surface using natural Canada balsam. The coverslip should be supported in the vertical position to the work table with a small amount of wax. When the wax and the balsam have hardened, the blade can be advanced and the second cut made (see Fig. 1.17). The wax support can be removed carefully with xylene, and the section left on the coverslip with a slice being mounted on the second surface. Alternatively the section can be removed using xylene and mounted in the normal way.

Saw Blade

Section Being Cut

Cover Slip

Natural Canada Balsam

---1+-+----- Specimen

FIG. 1.17. Using the annular saw to cut the section.

36 A. D. Curson

1.6 MELT PRESSINGS

Thin films of semicrystalline polymers can be produced by pressing small pieces between a slide and coverslip at selected melt temperatures and subsequently cooling under any chosen crystallisation conditions. Such preparations are useful for observing modifications to crystalline texture produced by different thermal treatments, and for the determination of particle size of additives, e.g. pigments. However, the method has some serious limitations:

- The dispersion of the phases in mUltiphase systems may be changed.

- Degradation may occur giving rise to non-representative crystalline texture.

- Agglomerates of additive in the original sample may be broken up and dispersed.

- The crystalline texture may not necessarily be that which would be developed in bulk under equivalent thermal conditions, owing mainly to surface nucleation and melt shear effects.

Despite these disadvantages, melt pressing is a quick and, therefore, popular technique. Except in a few specific applications, such as for depolarised light intensity analysis, or for the characterization of the growth of crystalline structure, pressings should not be thicker than about 30 f../m. With most polyolefins the resulting pressing can, if required, be removed from the slide and coverslip and sectioned as for a film. Nylon and PET pressings need to be separated from the glass surfaces using a thin, high-temperature resistant film, such as a polyimide, if sectioning of the pressing is to be undertaken.

1.7 STAINING

Staining to increase image contrast in light microscopy is not widely applicable in the synthetic polymer field, but three specific instances are worth noting:

(1) Unsaturated rubbers in sections can be selectively stained black using osmium tetroxide.

(2) The addition of a fluorochrome to the methyl methacrylate used for embedding purposes can be useful in demonstrating impregnation of the specimen.


(3) Voiding or porosity exposed in cut surfaces can be more easily seen and the extent of penetration established if the surface is impregnated with drawing ink. The technique is to deposit the ink on the surface, subject it to a vacuum cycling routine and then draw off the excess ink with a pipette or tissue. When the remaining ink is dry the surface is polished gently on a dry tissue or cloth, after which the inkfilled voids will be clearly visible.

1.8 FINAL COMMENTS

Good specimen preparation is the essential precursor of good microscopy. Because of the variability of polymeric materials, not just between types and grades but also between batches, and also the vagaries of processing techniques, each sample will be different from the next. Consequently there are no 'standard routines' or 'magic buttons' that would ensure good results. This chapter has not covered all possible techniques, or even all variations of those with which it does deal. The secret of success is a sure knowledge of what is required, an appreciation of how the sample will react to different procedures, the awareness and skill to perform as many varied preparation techniques as possible and to modify these as necessary, and patience.

2

Basic Light Microscopy and the Phase Contrast Microscope

D. A. HEMSLEY

Polymer Microscopy Services, Loughborough, UK

2.1 INTRODUCTION

The theory of the light microscope is well established and copious literature which examines the theoretical aspects of the instrument from a diversity of viewpoints is widely available. l -4 It is not intended that this chapter should again cover this well trodden ground in detail. However, it is felt necessary (Section 2.4) to draw attention to a number of theories of the microscope. One of these provides the background for the description of a modification of the microscope which increases image contrast and which is especially useful for the examination of polymeric specimens.

The contents of this book amply illustrate the diversity of microscopical techniques applicable to polymers. This chapter is confined largely to two basic techniques. The simpler is when the microscope is used purely as an aid to resolution, effectively extending the visual acuity of the user's eyes. Perhaps surprisingly, there seems no generally agreed phrase to describe such a microscope and to distinguish it from its more complex modifications such as the fluorescence, interference or phase contrast microscope. Terms describing the basic technique as 'bright field microscopy', 'ordinary light microscopy' or 'common light microscopy' seem to be interchangeable in the literature. The present author has a preference for the last of these but accepts that a logical justification for the term is difficult since there is nothing 'uncommon' about the light used in, for example, the phase contrast microscope.

39

40 D. A. Hemsley

As far as common light techniques are concerned, both transmitted and reflected light methods and their applications will be considered in this chapter, although the latter receives less emphasis. This reflects the current utilization pattern in both industry and academic research. The pattern stems from the relative ease with which polymers can be prepared as thin sections and their inherent optical transparency. The phase contrast techniques discussed in Sections 2.6 and 2.7 are now virtually confined to transmitted light applications. Although some reflected light phase contrast systems are still available, their use in recent years has fallen away almost totally in favour of the differential interference methods described in Chapter 5.

In transmission the choice between phase contrast and interference methods is much less obvious and the relative virtues and vices of these two systems need to be examined carefully.

2.2 SYNTHETIC POLYMERS AS SPECIMENS FOR LIGHT MICROSCOPY

Light falling upon a specimen will be affected in a number of ways. Although distinct in practical terms, it should be recognized that at the deeper level of optics the interactions between light and the specimen are united in a common theory. From the microscopist's standpoint, however, it is more helpful to consider these as separate and distinct processes, such as absorption, fluorescence, reflection, refraction and diffraction, especially when specialized forms of light microscope are used to capitalize on a specific interaction. For example, a microscope intended to obtain information about a specimen by the phenomenon of fluorescence will have design features greatly enhancing its ability to work efficiently in this particular mode.

The most usefullight/specimen interaction is absorption, since this readily provides optical contrast in the image produced by the microscope. However, most synthetic polymers show negligible absorption in the range of wavelengths representing the visible spectrum (roughly 400-700 nm). This is potentially a serious handicap to the microscopist since the visibility of structural detail in a specimen depends on intensity or colour contrast. Obviously the situation in respect of dyed or pigmented polymer compounds is different; here there may well be sufficient absorption to provide image contrast. If this absorption is wavelength selective, colour as well as intensity contrast is achieved. This raises the

Basic Light Microscopy and the Phase Contrast Microscope 41

question of whether synthetic polymer specimens can be selectively stained to increase image contrast. Staining is a standard method widely employed for biological materials, most of which are natural polymers. Although some useful degree of staining can be obtained when working with certain polymers, particularly the polyamides and acrylics, the processes necessary to ensure sufficient strain uptake are often inconvenient and likely to modify the structure of the material. The use of osmium tetroxide has for many years found favour with electron microscopists. It is used to great advantage for light and electron microscopy on polymers possessing a degree of molecular unsaturation, but the range of materials to which it can be applied is very limited. The net result is that staining methods are not widely employed, and in the light microscopy context they rightly receive only a cursory mention in Chapter 1.

Absorption outside the visible spectrum is both possible and useful as a contrasting mechanism. In particular, techniques have been used to explore the distribution and concentration of ultraviolet radiation absorbing additives. UV microscopy is discussed in detail in Chapter 7.

Microscopy beyond the visible spectrum into the longer wavelength region - infrared microscopy - suffers from a fundamental problem. Although contrast is obtainable from polymeric specimens by this method, the wavelengths that need to be employed militate against high spatial resolution. Nevertheless microscopes capable of working in the IR region have been available since around 1953. These early models were crude compared with modem instruments which are designed to obtain IR absorption spectra from selected parts of the microscope image rather than just produce image contrast.

In the absence of absorption, other methods of obtaining contrast in the image must be found. Fortunately for the polymer microscopist this problem has already received close attention in many other fields of application of the light microscope. Contrast enhancement by optical methods utilizes the refractive optical characteristics of the specimen. Perhaps the simplest example is the use of crossed polars to generate image contrast from the double refraction or 'birefringence' effects demonstrated by most crystallizing polymers. Indeed the use of the polarizing microscope to observe crystalline microstructure represents one of the earliest, and most widely known, methods of polymer microscopy. Useful contrast using polarized light may also be obtained from specimens showing molecular orientation, stress or form birefringence. In the last example image contrast is the result of periodicity in the

42 D. A. Hemsley

composition of the polymer and is of particular use in the study of certain block copolymers. Qualitative polarized light methods are discussed in Chapter 3, and some quantitative aspects in Chapter 4.

Polymer systems in which there are variations in refractive index due to the chemical composition, such as composites, represent a different class of specimen. If the refractive index differences between phases in the composite are large, say greater than 0'05, strong diffraction effects at the phase boundaries will enable these to be seen without difficulty. However, although the range of refractive indices for commercial polymers is wide (roughly 1·3 to greater than 1'7), many are centred around 1· 5, so phase separated polymer/polymer mixtures tend to show small refractive index fluctuations and the phase boundary diffraction effects are weak. Such polymer systems are best examined using microscopical techniques specially developed for 'phase objects' such as the phase contrast, differential interference contrast or Hoffmann modulation contrast methods described in this or following chapters. As examples one might cite Nylon/PTFE blends as a case where the refractive index difference is large (l. 53-1' 36) and common light observation will show the distribution of the two phases. On the other hand, a Nylon/polybutadiene blend (l'53-l'52) would demand the use of an optical contrast enhancement method.

Note that at present only the detection of the phases is being considered. Although the Becke line or Van der Kolk tests5 will allow determination of whether a particular phase has a higher or lower refractive index than its surroundings, no optical information permitting identification of the phases is present in the common light image. Both phase contrast and differential interference contrast can help in identification, but the most satisfactory approach is to use the transmitted light interferometric methods described in Chapter 6.

'Real' polymer compounds may well contain optically non-absorbing inorganic additives, in some cases at high loadings. The above discussion concerning polymer/polymer 'phase objects' is equally applicable to such compounds. Additionally, many inorganic additives are crystalline and highly doubly refracting, so polarized light methods of contrast acquisition are important, especially when the host polymer exhibits low or, better still, zero crystallinity.

From the specimen preparation point of view, polymers present a variety of challenges, and ways of meeting them are discussed in Chapter 1. Some polymers, such as polypropylene, PVC, polyacetal and polystyrene, are easily prepared unless they exhibit a very high degree of


molecular orientation. The difficulties are hardly more than would be encountered in thin sectioning biological tissue. Hard polymers, including polyimides, PEEK. UF and PF resins, are best treated as though they are metallic or geological specimens, and as such they present few problems for reflected light and transmitted light work respectively. The difficult polymers to prepare are those that are excessively soft or elastic. The problems associated with these materials, particularly if very thin sections are necessary, can be formidable but not necessarily impossible to overcome.

To summarize, synthetic polymer specimens can be prepared and microscopically examined by a variety of techniques. The microscopical methods used will often include those chosen for image contrast enhancement or for the quantitative determination of refractive index or birefringence. The results of the examination of a polymer product can be very rewarding in terms of in-service performance prediction, the optimization of production process variables and the identification of the cause of product failure.

2.3 LIGHT VERSUS ELECTRON MICROSCOPY OF POLYMERS

To a polymer technologist or scientist with a problem to solve, any division between light and electron microscopy is artificial. The microscopical examination of a specimen is usually embarked upon with clear aims in mind. It is the problem that should dictate the choice of method, and the examination will often, quite justifiably, involve the use of a range of microscopical techniques, including both light and electron microscopy.

The diversity of light-based techniques is surprisingly wide. In very general terms, light microscopes are cheaper than their electron counterparts, less expensive to run, and there are almost no problems of adverse interactions between the specimen and the radiation used. In the light microscope the specimen is not exposed to a vacuum and the specimen preparation procedures are fairly straightforward and flexible.

The great advantages offered by the electron microscope are substantially higher resolution in several modes of operation and, through accessories, elemental analysis of very small volumes. An increased depth of field is another advantage, at least in the case of the conven!ional scanning microscope. This allows easier image interpretation (see Fig. 2.1) and a more precise understanding of the spatial

44 D. A. Hemsley

interrelationships offeatures in the image. Inevitably, when working at the highest resolution, only an extremely small volume of material is being examined, the significance of which to the investigation in hand may be in doubt unless many fields of view are examined. Thus the benefits of high resolution may be at least partially offset by a massive expansion in the amount of work required to support or disprove any hypothesis.

In the general context of routine industrial problem solving, the transmitted light microscope finds more application than its electron counterpart. In the examination of polymer surfaces this situation tends

(a)

FIG. 2.1. The fracture surface of an acrylic specimen imaged using (a) scanning electron microscopy (secondary electron mode) and (b) reflected light microscopy. The image produced by the latter technique is inferior because of the smaller depth of field and lower resolution. Also, in the case of the tilted block-like central feature, light is being reflected outside the aperture of the objective lens, making it impossible to comment on the microstructure

(both X405).


(b)

FIG. 2.1.-contd.

to be reversed. The much greater depth offield of the scanning electron microscope, and the fact that small routine instruments are now relatively cheap, means that these are rightly often the prefered tool for examining rough surfaces, even at magnifications comparable to those possible with the light microscope.

In no sense is the light microscope the poor relation of the electron microscope, neither is the latter to be considered by the light specialist only when all else fails. It is the author's experience that two basic misunderstandings seem to persist in many research and development laboratories. First there is the view that, because of its intrinsically higher resolution (and cost), electron microscopy must always be the most appropriate technique. Second, because the light microscope has been with us since the 17th century, its day is past! Both of these views are untenable in practice. In materials science and technology in general, and it can be argued in the case of synthetic polymers

46 D. A. Hemsley

especially, both techniques are important tools to be used as and when appropriate. One hesitates to insist that any examination with an electron microscope should be preceded by light microscopy, but polymer science and technology would benefit if this were a more frequent procedure.

2.4 BASIC LIGHT MICROSCOPY

2.4.1 Image Formation It has already been stressed that it is not intended to discuss in detail the basic light microscope or its operation. References 1-4 should provide enough background for the prospective microscopist to use the instrument with sufficient consideration and care to avoid major practical problems.

It is unfortunate but undeniable that most microscopes will provide some kind of image despite being far from their optimum adjustment. At best a poorly adjusted instrument will reveal little of the true nature of the specimen. At worst 'structural detail' will be seen in the image which is absent from the polymer sample being examined. The need for discipline in correctly adjusting a microscope to give the best image of which it is capable is therefore paramount, and more than a passing glance at the recommended texts will pay dividends in this respect.

An understanding of the basic theory behind the operation of a microscope helps with achieving correct adjustment and also with subsequent image interpretation. Microscope theory can be approached from several different directions. In the literature the choice is usually made according to the likely background of the reader and whether the basic theory will subsequently need to be developed to explain the working of a more advanced type of microscope.

Perhaps the most obvious and simplest way of describing the function of the components of a transmitted light microscope is through the use of a diagram such as that in Fig. 2.2 which displays the relative positions of the components and shows how rays oflight progress from the lamp filament to the retina of the user's eye. The shortcomings of the ray path approach become most noticeable when dealing with the function and performance of the objective lens, in particular with its limitations in resolving fine detail. A more satisfactory approach is due to Ernst Abbe who, in the late 19th century, produced a theory of image formation based directly on diffraction theory.


] EYEPIECE

s

A--~n-:::'--J---

F---

FIG. 2.2. Path of the imaging rays through a standard transmitted light microscope: L, lamp; F, field iris diaphragm; A, aperture iris diaphragm;

S, specimen; PIP, primary image plane; E, eye.

Consider the case in which a diffraction grating (a two-dimensional periodic structure consisting of alternate opaque and transparent strips) is to be imaged into the primary image plane of the microscope. This is the task of the objective lens. Suppose also that the grating is illuminated by a narrow collimated beam of light from the condenser unit as shown in Fig. 2.3. As a result of the diffraction phenomenon, light that has passed through the grating will be redistributed to give a series of intensity maxima. One of these will be in the 'straight through' direction and is referred to as the zero-order beam. Others will occur at angles al.a2 to this direction on either side of the zero-order beam. If the

48 D. A. Hemsley

2 o 2

F--- ---F

o -+-T+-+--i---i-+-+-+--J-- 0

FIG. 2.3. Diffraction of light by a grating: G, grating; CC, condenser lens; 00, objective lens; FF, back focal plane of objective lens; 0, I and 2 refer to the

diffraction order.

geometrical diameter of the aperture of the objective lens and its distance from the grating are such that this diffracted light can enter the lens, the beams are brought to focus in its back focal plane. If this plane is viewed it is possible to see a set of 'diffraction spots' as shown in Fig. 2.4. Note that the focal plane displays the diffraction angles (a) as distances measured outwards from the central zero-order spot position.

Light from the diffraction spots passes on through the microscope to the primary image plane. Here the phase relationship between the light waves is such that optical interference takes place to give a pattern of intensity which is the image of the object.

Two important conclusions follow from Abbe's approach. First, it is clear that to form an image more than one beam must reach the primary image plane. Interference cannot occur and no image is formed if only one beam reaches this plane. Such a situation would occur if the diffracted beams in Fig. 2.3 fell outside the aperture of the objective lens.

The ability of this lens to collect over a wide range of grating diffraction angles can be expressed in terms of its numerical aperture


FIG. 2.4. Diffraction spots produced by a linear grating and viewed in the back focal plane of the objective using quasi-monochromatic light. Note that, having adjusted the microscope for Kohler illumination, an enlarged image of the lamp

filament appears within each diffraction spot.

(NA) which is given by n sin e, where n is the refractive index of the medium between the lens and the grating and e is the angle subtended by the radius of the lens aperture as shown in Fig. 2.5. Thus the higher the NA the greater the ability of the lens to collect the diffracted beams. Since the grating diffraction angles are an inverse function of the grating spacing, a high NA lens will be necessary iflines on a grating with a short periodicity are to be resolved in an image.

A second conclusion might be that to produce the best possible image it is necessary to collect all the diffracted light from the grating. In practice the intensity of the diffracted beams falls off rapidly as the diffraction angle increases, so the omission of the waves diffracted at large angles has little effect upon the image in the primary image plane. Indeed a serviceable image can be obtained by interfering only one diffracted beam with the zero-order beam.

The Abbe theory outlined above explains many aspects of objective lens performance. For example, since the diffraction grating will diffract red light through a larger angle than blue, it would be expected that the limit of resolution for blue light would be lower (i.e. better) than for red; this proves to be the case in practice. This does not mean that all polymer specimens should be examined using a blue filter in the light

50 D. A. Hemsley

FIG. 2.5. Definition of numerical aperture: NA = n sin e, n being the refractive index of the medium between the specimen and the

objective lens.

path. This is generally unnecessary but, in situations where the highest possible resolution is desired, the marginal gain by doing this can be justified.

Two objections to the theory as expressed above are clear. Polymer microscopists do not normally spend time looking at diffraction gratings (although this is recommended as an instructive exercise for the newcomer to microscopy), and the use of a narrow collimated beam to illuminate the specimen is not standard practice. The second objection is answered by considering an arrangement in which the specimen is illuminated by a cone of rays emerging from the condenser unit. The actual range of angles over which the objective can collect diffracted waves is now increased. In effect this means that it is the NA of the condenser/objective combination that is of importance in determining the resolving power of the microscope as a whole. In theory this is maximized when the NA of the condenser matches that of the objective lens. In practice the condenser NA is kept somewhat smaller, especially when using crossed polars to observe the crystalline texture of the polymers (see Chapter 3).

The effective condenser NA is controlled by the diameter of the aperture iris in the front focal plane of the condenser lens. The setting of this diameter forms part of the standard setting-up procedure for Kohler illumination described elsewhere.2 Failure to optimize this setting has an adverse effect upon image quality.

An answer to the first objection relies on the fact that a non-periodic object can be regarded as a set of periodic objects added together.4

A third approach to microscopy theory, closely related to the above concepts, uses some of the ideas of information theory. This sees the specimen as composed of a set of spatial frequencies.6 A diffraction


grating which is sinusoidal in optical density would be regarded as presenting a single spatial frequency. On the other hand, a more complicated distribution of optical density, such as presented by a 'real' specimen on the microscope stage, can be resolved, by Fourier analysis, into a spectrum of spatial frequencies. One of the functions of the microscope is to transfer this spectrum from the object to the image plane. It can be regarded as a filter, in the electronics sense, with specific transfer characteristics that in reality are less than ideal. For high resolving power the system must be able to pass high spatial frequencies, but in addition the opportunity exists to 'process' the 'signals' to enhance image contrast. This approach has appeal when considering the behaviour of various optical methods of contrast generation and control, and links with the formal assessment of lens characteristics obtained by measurement of optical transfer functions.

2.4.2 Practical Considerations The special instrumental requirements for the practical microscopy of plastics and rubbers are few. Mostly these are discussed below (phase contrast microscopy) or in the following chapters.

However, two points deserve comment. First, many of the special techniques are inefficient in terms oflight transfer. The problem of the resulting low image brightness is compounded by the characteristics of many polymer specimens. Often they exhibit very low birefringence between crossed polars, and in reflected light problems arise because of the low reflectivity of polymers ( -4%). The microscope therefore needs to be fitted with a high intensity light source - typically a low voltage 100 watt tungsten halogen lamp.

Some techniques require the user to observe the back focal plane of the objective lens (e.g. small-angle light scattering and conoscopy), and it is useful to be able to insert a ground glass screen into the illumination system. This diffuses the image of the lamp filament which should occur in this plane if the microscope has been correctly adjusted for Kohler illumination.

The routine light microscopy of plastics and rubber seldom requires the instrument to be used at its limits of resolution. Indeed, an especially low power X 1 objective lens may be of more value in the objective set than a X 100. If the use of higher magnifications is necessary it is particularly important with polymeric specimens that the user commences the observation at low power and works up. The interrelationships between microstructural features in polymers can be

52 D. A. Hemsley

complex, and isolated observations at high magnification can be misleading.

Finally, it is a characteristic of polymer work that several methods of examination may be necessary to obtain as complete as possible a picture of the microstructure. It is therefore an advantage if the microscope stand will accept a range of accessories for contrast enhancement, quantitative measurement and photomicrography.

2.5 APPLICATIONS FOR COMMON LIGHT MICROSCOPY

2.5.1 Observations on Pigment and Other Particulate Additives As discussed earlier in this chapter, most polymers may be classified as 'phase objects' since few show any appreciable intrinsic absorption in the visible spectrum. On the other hand, commercial polymer systems are almost invariably pigmented, and may contain other additives such as fillers or particulate stabilizers.

Observation of the dispersion and distribution of such particulate additives is one of the main applications for common light microscopy in both qualitative and quantitative modes of operation. A poor distribution of additives may have dire effects upon the in-service survival of a manufactured product. Not only will the properties that the additive is intended to convey to the polymer (e.g. colour or stabilization) be locally variable, but the presence of undispersed 'agglomerates' of particles may seriously and adversely affect the strength of a product by the provision of sites of high stress concentration when loaded.

Additives are often introduced into polymer systems by a 'masterbatch' technique. Masterbatches consist of a high concentration of additives (typically 30%) distributed in polymer. This is subsequently 'let down' in 'natural' polymer to give a typical final concentration of around 1 %. The mixing of the masterbatch into the natural polymer usually takes place during the production, by extrusion or injection moulding, of the final manufactured article. It is possible to examine the additive within a masterbatch by transmitted common light microscopy but the microtomed section thickness required is very small, often less than 1 pm. This even applies to masterbatches containing carbon black. However, such materials should be examined with close attention to possible heating of the specimen by the light beam. The high absorption of the carbon over a wide range of wavelengths produces significant heating to the detriment of the surrounding polymer. This phenomenon is one of


the more spectacular examples of 'beam damage' in the light microscope - a problem more familiar to the electron microscopist. The use of a heat-absorbing filter in the illumination system (always good practice) reduces the problem, but extra filters of this type, or even colour filters, may be necessary to avoid difficulties with more sensitive specimens such as high loadings of carbon black in low density polyethylene.

Historically the distribution and dispersion of carbon black in rubber has been 'quantified' on a routine basis by comparison of the microscopic image with a set of standard photomicrographs. The Cabot carbon tese is an example which is still widely employed. Obviously such subjective tests, which have been extended to include other polymers such as polyethylene [BS 2782: Part II; method l106A (1983) and methods 823A,B (1978»), must be carried out at a specified microscope magnification. Despite specimen preparation difficulties, such tests remain popular because they require little by way of equipment beyond the common light microscope itself.

More generally there appears to be promising scope for new image analysis techniques in this particular area of polymer microscopy. A variety of procedures for pigment distribution analysis have already been devised8. 9 to yield data useful in both product quality control and more fundamental studies of the effect of pigment distribution on properties and on the efficiency of mixing and compounding equipment. Microscopically it is necessary only to obtain a sufficiently 'dilute' image of the pigment particles by choosing an appropriate section thickness. In theory at least, analysis is then straightforward.

Typical examples of a well dispersed and a poorly dispersed carbon black in a rubber are shown in Fig. 2.6. Although the concentration of the additive in the polymer is high, it is still possible to obtain a section thin enough (1 .um) to identify undispersed material. Such images can be quantified by modern electronic image analysis without much difficulty, thus assisting correlation between dispersion and mechanical or polymeric properties.

The specimen preparation step, particularly with reference to carbon black in rubber, can be simplified for routine use by using reflected rather than transmitted light methods of image formation. A typical method has been described by Mutagahywa and Hemsley.lO Such methods are intended more for routine comparison of specimens than for obtaining the fundamental parameters describing the distribution and dispersion of the additive.

54 D. A. Hemsley

(a)

FIG. 2.6. (a) Well dispersed and (b) poorly dispersed carbon black in a thin section of rubber tyre compound; transmitted light (X 180).

Pigment agglomeration or pigment streaking are in practice a major source of product failure. Just how bad things can be is shown in Fig. 2.7. As already mentioned, by producing stress concentrations in the product when it is loaded, such a poor distribution of pigment can contribute to mechanical failure regardless of any optical effects involved. An inappropriate selection of masterbatch or inadequate processing of the compound is usually to blame.

2.5.2 Detection and Identification of Contaminants Contamination of manufactured plastics and rubber products is not uncommon and the sources of contaminants are many and varied. There are three main types of contamination:

- Contamination of the raw material


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- Contamination by processing machinery - Contamination by the environment.

Raw material contamination is most often polymeric, although inadvertent inclusion of traces of 'foreign' pigments, fillers or other additives may also occur. Polymer/polymer contamination may be difficult to see using common light microscopy unless the refractive index difference between the polymers is large. Other techniques, such as polarized light or phase contrast microscopy, may be more appropriate in such cases. On the other hand, particulate materials such as pigments, reinforcement fibres or fillers are usually more visible, though not always. Glass fibres in Nylon can be difficult to see, as can silica in polyethylene.

Contamination introduced during processing often includes such materials as metals, rubber or oil. Of these, metals will be clearly visible using common light microscopy, and it is likely that any rubber will

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itself contain particulate additives. The recognition of oil contamination is more difficult and if the presence of mineral oil is suspected the specimen is best examined using an ultraviolet autofluoresence technique (see Chapter 7). Severely degraded polymer, usually reddish brown or black in colour, is readily detected but less easily positively identified. A clue is that the boundary of the contamination will be iUdefined if the degree of degradation is not severe and if the material is the same as the host polymer. Again degradation may be accompanied by the development of UV autofluorescence, especially in PVC compounds, and this can be a valuable additional observational and confirmational technique if the degree of degradation is modest.

Environmental contamination is the most varied of all, ranging from fibres (including hair, asbestos and paper), rust and paint flakes through to building materials such as sand, brick dust and cement. Airborne contamination by pollen, seeds or even whole insects may be found in plastics products, although usually considerably modified by the manufacturing process!

Positive identification of contamination usually requires a combination of microscopical methods, and it is often surprising how readily common contaminants can be identified on the basis of their shape, colour, refractive index contrast, birefringence and other optical properties such as dispersion. Spot chemical tests can be useful although they appear less popular than they used to be. Mason ll gives details of some of these tests and a reference list. In some cases a micro hot-stage can be of value for determining crystalline melting points.

A valuable general aid to contamination identification is the Particle Atlas. 12 More specifically, the book by Winchell and Winchell13 helps in the identification of inorganic contaminants from their optical characteristics. Some excellent fibre identification schemes have also appeared in the literature.14

2.5.3 Examination of Surfaces The results of an examination of a polymer surface by common reflected light microscopy are often disappointing. Very rough surfaces, such as those presented by ductile fractures or textured plastics products, are most profitably examined using a scanning electron microscope (SEM). Very smooth surfaces, of which the natural surface of films and the 'mirror' region of brittle fractures are good examples, are best examined by one ofthe interference techniques discussed in Chapter 5. Surfaces of intermediate roughness can sometimes be examined successfully by

58 D. A. Hemsley

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common reflected light but the interpretation may be difficult and the problems are compounded by the low surface reflectivity of polymers. The latter problem may be overcome by metal coating of the surface as described in Chapter 1.

The contrast in images of surfaces that diffusely scatter a high proportion of incident light may be considerably improved by reducing the diameter of the field iris of the microscope. This does not affect the resolving power, although the illuminated field of view is reduced. A substantial improvement in contrast can be obtained in the image of a surface of a polymer containing a strongly scattering pigment such as titanium dioxide. This effect is shown in Fig. 2.8.

2.6 PHASE CONTRAST MICROSCOPY

Many polymer specimens, provided that they are free of dyes or particulate additives, absorb little or no light. As indicated in Section 2.2, these specimens are termed 'phase objects' as opposed to 'amplitude objects'. However, there may be sufficient fluctuation in refractive index within the specimen to give rise to diffraction, scattering or refraction effects. These allow the edges of structural features to be seen in the common light microscope. More usually the refractive index fluctuations are small and the visibility of features is difficult, if not impossible. It was for this type of specimen that the phase contrast microscope was developed by Zernicke in the 1930s. Although phase contrast can be employed in either a transmitted or a reflected light mode, its use on polymers as a reflection technique is rare. For this reason it is discussed below only in terms of transmitted light.

2.6.1 Basic Principles In Section 2.4.1, in outlining the Abbe theory of image formation, it was observed that the primary image was formed by optical interference of the waves emerging from the back focal plane of the objective lens of the microscope. In considering the relatively simple case of an object consisting of a linear grating of opaque and transparent stripes, discussion was confined to an 'amplitude object'. Furthermore there was no discussion of the relative phase of the waves taking part in the interference process. In fact there is approximately a half-wavelength (180°) phase difference between the waves emerging from the zeroorder diffraction spot and those from the higher orders. A number of


authors have described the origin of this effect mathematically and more graphically by the use of vector diagrams.4• 15

It can be shown that, if an amplitude grating is replaced by another consisting of grooves cut in a transparent sheet (a 'phase grating'), diffraction effects in the back focal plane of the objective are still observed. However, it can also be shown that the phase difference referred to above is reduced to approximately one-quarter of a wavelength (90°) provided that the optical path length differences in the grating are small. In essence the phase contrast method involves modifying the phase relationship between the diffracted and undiffracted light so that a more favourable situation exists in terms of image visibility. Specifically the aim is to increase the phase difference from around 90° to around 180°. This can be accomplished by allowing the undiffracted light to pass through a thin plate (a 'phase plate,) of such a thickness that the necessary phase shift is produced. The obvious place to position the phase plate is in the back focal plane of the objective, because here the zero-order diffracted light can be located separately from the diffracted light, but to do this it is necessary to restrict the illumination cone from the condenser. Failure to do this would mean that zero-order beams from the range of illumination angles would cover the back focal plane. One answer might be to restrict the cone by closing down the aperture iris to give a thin collimated beam. The zeroorder diffraction maximum would then be clearly located at the centre of the back focal plane, but this would be accompanied by diminished resolution and a severe loss of light. It is therefore common practice to replace the aperture iris with an annular clear ring. The undeviated or zero-order light then passes through a well defined ring in the back focal plane of the objective. It is here that an annular phase plate is positioned. The system is shown in outline in Fig. 2.9. In practice two refinements to the system are needed. First, image contrast can be further improved if the amplitude as well as the phase of the zero-order light is adjusted. This involves making the phase plate both partially absorbing as well as phase shifting. Second, since the phase plate needs to produce a quarter wavelength shift of phase, this wavelength must be defined and utilized when the system is used. Thus it is not unusual to incorporate a suitable colour filter into the microscope. The bandwidth of the filter used is a compromise between the need to define closely the wavelength and the need to preserve an acceptable level of image brightness.

In practice phase contrast systems may be 'positive' or 'negative'. For

62 D. A. Hemsley

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the former, higher refractive indices in the specimen will show as being darker than the surrounding field; in negative phase contrast the situation is reversed. However, the contrast actually seen will also depend on the magnitude of the phase shift (or optical path differences) in the specimen. If these are no longer small, as usually assumed by the theory, contrast will be modified or even reversed. A treatment of the theory by Goldstein l6 is not restricted to specimens showing small phase shifts.

2.6.2 Shortcomings of the Phase Contrast Method It has already been mentioned that the phase contrast system is inefficient in terms of light usage. This is of more importance when the system is used for photomicrography rather than for a direct visual examination of the specimen.

Discussion of the characteristics of the specimen has concentrated on refractive index, but specimen thickness also influences the magnitude of optical path differences in the specimen (see Chapter 6).


In practice many polymer specimens are examined as thin sections which may exhibit knife marks. Since these represent regions where the specimen is thicker or thinner, the actual phase shifts produced by the specimen will be locally variable and may no longer be small. The result is that any knife marks may be made substantially more visible by the phase contrast method, placing greater demands on section quality.

A characteristic of the phase contrast image is the 'halo' effect, illustrated in Fig. 2.10. This can considerably complicate image interpretation, particularly if the field of view shows a concentration of features. According to Spencer,4 the origin of the halo is the proportion of diffracted light that passes through the phase ring intended to pass

FIG. 2.10. Halo effect around the periphery of a PVC powder particle; transmitted light (X400).

64 D. A. Hemsley

only zero-order beams. The halo is therefore an unavoidable characteristic of phase contrast systems.

One clear advantage ofthe system is its great sensitivity. It compares with, and often betters, the interference systems described in Chapters 5 and 6. Furthermore the system is relatively inexpensive compared with the interference equipment.

Another advantage, perhaps less obvious, is that the phase contrast system does not involve the use of polarized light. Almost all the differential interference methods currently available for image contrast generation employ devices whose operation depend on the input light being plane polarized. This can cause serious interpretational difficulties with polymers; these are discussed at greater length in Chapter 5. The problems arise because many polymeric specimens, or phases within them, are to some degree optically anisotropic and therefore not characterized by a single refractive index.

2.6.3 Adjustments and Alignment of a Phase Contrast Microscope A microscope for phase contrast work requires a special condenser unit to provide a set of illuminate annuli, a set of objectives containing the necessary phase rings, and a method of observing the back focal plane of the objective.

As always, the correct adjustment and alignment of the microscope is essential if satisfactory results are to be forthcoming. The practical procedure can be summarized as:

(1) Observe the specimen with the desired objective and select the necessary corresponding annulus in the condenser. The objective/ annulus pairs are usually coded by the manufacturer.

(2) Set up Kohler illumination in the usual way.2 Note that the aperture iris adjustment is no longer part of the setting up procedure since this has been replaced by an annulus.

(3) Observe the back focal plane of the objective. There are several ways of doing this. One method is to replace an eyepiece of the microscope by a focussing phase telescope. In the absence of such a device the back focal plane can be seen by looking directly down the tube from which the eyepiece has been removed. The image obtained is then smaller than with the telescope, and subsequent adjustment is more difficult. A third possibility, if the instrument is also used for polarized light microscopy (a common situation in polymer work), is that it may be equipped with a Bertrand lens. When inserted (and


focussed if the adjustment is provided) it works together with the eyepiece( s) to view the objective back focal plane.

(4) In the back focal plane are images of the condenser annulus (bright) and the phase ring of the objective (dark). Using the annulus centring controls, these two images should be made to coincide. If the correct selection of annulus has been made, its image should be totally covered by the image of the phase ring.

(5) Remove the telescope or Bertrand lens and, if necessary, replace the eyepiece for normal observation.

2.7 APPLICATIONS FOR PHASE CONTRAST MICROSCOPY

The principal area of application is in the study of multiphase polymer systems. Obviously the phases must be large enough to be resolved by the light microscope, but once that condition is satisfied the system will normally provide a satisfactory high contrast image. Typically the system finds application in looking at materials toughened by the addition of a rubbery phase such as HIP (high impact polystyrene), ABS (acrylonitrile-butadiene-styrene terpolymers), or ABS modified polycarbonate (see Fig. 2.11). For maximum contrast it is usually necessary to examine these materials at section thicknesses of 2 f.lm or less. The presence of particulate additives of high refractive index may severely inhibit the interpretation of a phase contrast image because of the many overlapping 'halos' produced.

The layers in coextruded or laminated products may also be investigated using this technique. For example, the small refractive index difference between low density polyethylene and a copolymer of ethylene and vinyl acetate may cause difficulty in characterizing layers in certain coextruded packaging films. The phase contrast method allows adequate contrast to be achieved. However, when examining coextrusions or laminates of unknown construction, care is necessary not to interpret the halo along layer boundaries as a thin additional layer in the product.

Although the spherulitic texture of crystallizing plastic is conventionally examined in the polarizing microscope, the type of image obtained using phase contrast, such as that shown in Fig. 2.12, can be more informative if the fibrillar sub-structure is of particular interest. Some workers have used phase contrast and crossed polars simultaneously in examining specimens of this type. The interpretational

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difficulties that arise make it questionable whether this combined technique is worthwhile.

Cross contamination between polymers is also conventionally investigated using phase contrast microscopy. Although it is not easy to use the method to obtain precise data on the refractive index of any contamination, and hence to identify it, some idea of the relative indices of the phases present can usually be gained.

2.8 DARK GROUND MICROSCOPY

In principle this is an extension of the phase contrast method; a good account has been given by McLaughlin. 17 Instead of just modifying the relative phase of the zero-order light relative to the diffracted light, the former is removed completely. The Abbe theory indicates that an image can still be formed but contrast is reversed, with diffracting features in the object appearing in the image as bright upon a dark, ideally black, background.

The dark ground system is of value in polymer work both in its transmitted light and reflected light forms. The simplest procedure for preventing undiffracted light from contributing to image formulation would be to place a suitable opaque stop in the back focal plane of the objective or indeed in the filter carrier of the condenser unit (in transmitted light). Although such methods are reasonably effective at low powers, the contrast obtained is usually poor.

For critical transmitted light work a dark ground condenser should be used. This illuminates the specimen with a hollow cone of light which, if not redistributed by a specimen, falls outside the aperture of the objective lens. Once a diffracting specimen is placed on the microscope stage some light will now enter the objective and contribute to image formation. The amount oflight is small and as a result problems can arise with photomicrography. Nevertheless the technique is excellent for detecting small diffracting or scattering centres in polymer preparations. It is well suited to the examination of microvoiding or stress whitening as well as pigments and other particulate additives.

The reflected light mode operates in a similar manner but here light is usually led down around the outside of the objective lens and then deflected at an angle on to the specimen. Light reflected specularly from the surface of the specimen again falls outside the aperture of the

Basic Light Microscopy and the Phase Contrast Microscope 7l

objective lens. Diffracting features on the surface of the specimen, or slightly sub-surface if the polymer is transparent, then contribute to image formation as in the transmitted light version of the technique.

An impressive polymer application for the reflected light system is the observation of sub-surface light scatterers such as microvoids and pigments. If the surface of the polymer specimen is smooth, light reflected from it does not enter the objective. This 'unwanted' light is therefore not available to degrade the contrast of sub-surface features which are rendered much more visible as shown in Fig. 2.13. Furthermore the colour of pigment is readily seen against the dark background, and the technique has special value when studying plastic products in which there is a need to distinguish between pigment particles of different colours in the same specimen.

REFERENCES

1. Slayter, E. M., Optical Methods in Biology. Wiley-Interscience, New York, 1970.

2. Hartley, W. G., Hartley's Microscopy. Senecio Publishing Company, Oxford, 1970.

3. Martin, L. C, The Theory of the Microscope. Blackie, London, 1966. 4. Spencer, M., Fundamentals of Light Microscopy. Cambridge University

Press, Cambridge, 1982. 5. Hartshorne, N. H. & Stuart, A, Crystals and the Polarizing Microscope, 4th

edn. Arnold, London, 1970. 6. Lipson, H., Optical Transforms. Academic Press, London, 1972. 7. Medalia, A I. & Walker, D. F., Technical RG-124, 2nd edn. Cabot

Corporation Carbon Black Division, Boston, MA 1970. 8. Ess, 1. W. & Hornsby, P. R, Polymer Testing, 6 (1986) 205-18. 9. Ess, J. W., Hornsby, P. R, Lim, S. Y. & Bevis, M. 1., Plastics and Rubber

Processing and Applications, 4 (1984) 7-14. 10. Mutagahywa, B. & Hemsley, D. A, Plastics and Rubber Processing and

Applications, 5 (1985) 219-27. 11. Mason, C W.,Handbook of Chemical Microscopy, Vol. 1, 4th edn. Wiley, New

York, 1983. 12. McCrone, W. M. et al., The Particle Atlas, 2nd edn. Ann Arbor Science

Publishers, Michigan, 1979. 13. Winchell, A N. & Winchell, H., The Microscopical Characters of Artificial

Inorganic Solid Substances: Optical Properties of Artificial Minerals. Academic Press, New York, 1964.

14. Identification of Textile Materials, 7th edn. The Textile Institute of Manchester, 1975.

72 D. A. Hemsley

15. Home-Dickson, 1. (ed.), Optical Instruments and Techniques. Oriel Press, Newcastle upon Tyne, 1970.

16. Goldstein, D. 1., Journal of Microscopy, 128(1) (1982) 33-47. 17. McLaughlin, R. B., Special Methods in Light Microscopy. Microscope

Publications, London, 1982, p. 87.

3

Polarized Light: Theory and Measurements

B. P . SAVILLE

Institute of Polymer Technology. University of Technology. Loughborough. UK

3.1 INTRODUCTION

We are concerned in this chapter with the interaction of polarized light with matter in the form of dielectric materials such as polymers, and in particular with the phenomena that are observed when these materials are examined between crossed polars. In order to interpret correctly what is revealed when a specimen is examined under these conditions, it is necessary to understand more of the theoretical background than is the case in other branches of microscopy. For instance, there are four separate reasons why a specimen or part of a specimen may appear dark between crossed polars. Distinguishing between them is a matter of applying simple observational tests such as rotation of the specimen, but failure to do so will result in interpretations that may be diametrically opposed to the true ones. However, the information that can be gained about the molecular structure and orientation cannot in many cases be obtained by other methods.

3.2 LIGHT AND ITS INTERACTION WITH MATTER

3.2.1 Some General Comments For many purposes it is sufficient to consider light as some form of wave motion that is propagated with an extremely high velocity. It is not necessary in these cases to make any assumption whatsoever as to the type of displacement that takes place during the passage of the waves

73

74 B. P Saville

through a material medium. However, when the interaction oflight with matter is studied in order to deduce information about the structure of the matter, it is important to consider in detail what form this displacement takes, in terms of the physical construction of matter.

It is assumed, for the purposes of this chapter, that the molecules comprising a dielectric can be represented by bound charges which, after displacement by an electric field, are restored to their centres of equilibrium by an elastic force proportional to the original displacement. Light is an electromagnetic wave having a certain frequency range, one component of which is an oscillating electric field which is propagated in the direction of travel of the wave. Therefore, when an electromagnetic wave interacts with such a bound charge, it causes it to oscillate about its centre of equilibrium as the field reverses, and thus to act as a source of radiation itself.

If the frequency of the incident wave is the same as the natural frequency of vibration of the bound charge, there is resonance and a large proportion of the energy from the wave is given to the vibrating charge. In solid materials there is strong intermolecular action which causes this energy to be dissipated within the material, leading to strong absorption oflight at this particular frequency. In a gas at low pressure where there is little intermolecular attraction, the same phenomenon gives rise to strong radiation at this frequency. This effect is known as resonance radiation.

If the frequency of the impinging wave is not equal to that of the bound charge, the charge executes a forced oscillation of small amplitude, so the radiation from it is weak and of the same frequency as the electromagnetic wave. The phase of this secondary radiation is, however, different between the impressed frequency and the natural frequency of the bound charge. These secondary waves destructively interfere in all directions except the forward direction in which the original wave was travelling. They can therefore interfere with the primary wave and thus modify its phase. This phase change is equivalent to a change in the wave velocity of the original wave, since this is defined as the rate at which a condition of equal phase is propagated through a medium.

This is the mechanism whereby the velocity of light is reduced in media other than a vacuum. The interaction with the electromagnetic wave, and hence the reduction in the wave velocity, is dependent on the polarizability of the bonding within the molecule. Therefore, in structures where the bonds between atoms are not symmetrically arranged, it would be expected that the interaction of the material with

Polarized Light: Theory and Measurements 75

electromagnetic waves, as the light passed through the material, would depend on the direction of vibration of the electric field with respect to the bonding.

The refractive index is a measure of the velocity oflight in a material relative to its velocity in a vacuum. The higher the refractive index of a material the lower the velocity of light in it. It is possible to define an optical path length for light travelling through a section of a material as being the distance the wave front would travel in a vacuum in the same time that it takes to pass through the section. It is equal to the actual path length in the material multiplied by its refractive index.

In materials with non-symmetrical structures it is found that there is indeed a variation in refractive index with the direction of vibration of the electric vector of the light passing through it. This becomes apparent when the refractive index is measured using polarized light in which the vibration is confined to one direction perpendicular to the direction of propagation of the light.

Materials with a structure that is symmetrical in all three dimensions can be characterized by one refractive index and are said to be isotropic. Materials that need more than one refractive index to characterize them are said to be anisotropic. A structure that is completely asymmetric can be completely characterized by three principal refractive indices, in which case the material is said to be biaxial. Structures having two principal refractive indices generally have symmetry around an axis; they are known as uniaxial materials.

The terms uniaxial and biaxial refer to the number of optic axes in the material. An optic axis is a direction through the material for which light travelling in that direction would encounter the same refractive index regardless of its direction of polarization. The optic axis coincides with the axis of symmetry in uniaxial materials.

3.2.2 Atomic Polarization The effect of an incident electromagnetic wave on a non-metallic material is to cause polarization of the electric charges in the constituent molecules. That is, the positive and negative charges in the molecules, which previously balanced one another, are separated by the electric field, producing an electric dipole moment in the material. The dipole moment is the product of the separation of the two equal charges and their size. The dipole moment per unit volume of the material that results is known as the electric polarization, P. This is related to the applied field, E, by the expression

P = aE (3.l)

76 B. P. Saville

where a is known as the polarizability and is characteristic of the individual atoms involved modified by the bonding between them. On removal of the applied field the charges will revert to their former positions.

Polarization of the molecules in a material can take place by one of three mechanisms depending on the type of bonding present and the dipole moment of the original molecules.

1. In polar molecules which already have a permanent dipole moment, the dipoles will align themselves with the applied field. This is known as orientational polarization.

2. In non-polar molecules the negatively charged electron cloud can be shifted relative to the positively charged nucleus by the applied field, producing a spatial imbalance in the charge distribution. This is known as electronic polarization.

3. In ionic materials dipole moments are produced by a shift of the positive and negative ions with respect to one another under the influence of the applied field.

If the dielectric is subjected to a regular alternating field, as occurs during the passage of an electromagnetic wave through the material, the induced dipoles in it will try to follow the fluctuations in the electric field. Their ability to follow the field fluctuations at optical frequencies is determined by the moment of inertia of the system. For instance, in polar molecules the whole molecule has to rotate to follow the field reversals, whereas in non-polar molecules only the electrons, which have low inertia, have to follow the field fluctuations. Therefore the electric polarization produced in a molecule by electromagnetic radiation is dependent on the frequency ofthe applied field and the type of bonding present. For the frequencies involved in the visible region of the spectrum electronic polarization becomes the dominant mechanism.

It follows from this that in a molecule whose bonding is not symmetrical the polarization produced by an electric field will depend on the orientation ofthe electric field with respect to the different bonds. Polymer molecules have very directional bonding patterns; the atoms along the length of the chain are linked to one another covalently, but the lateral bonds between the chains are either non-existent or are weak hydrogen bonds or van der Waals forces. This means that, for an electromagnetic wave whose electric vector is vibrating parallel to the chain, the polarizability of the molecules is in general different from that for a wave whose electric vector is vibrating perpendicular to the chain.


The polarizability of a molecule is related to its refractive index by the Lorentz-Lorenz relation:

n 2 - 1 M 131TNa = R

n2 + 2 d - (3.2)

where n is the refractive index, d is the density, M is the molecular weight, N is the Avogadro number, a is the polarizability and R is the molecular or molar refractivity. The molar refractivities of molecules are additive, so the refractive index of a mixture can be obtained by summing the individual molar refractivities according to the numbers of molecules of each species present. It is possible using the principle of additivity of refractivities to assign a polarizability to each bond in a molecule by studying the refractivities of chain compounds. For example, the refractivity of the carbon-hydrogen bond is taken to be one-quarter of the molecular refractivity of methane. By using these methods, with certain simplifying assumptions, the main refractive indices of a polymer chain can be calculated theoretically. I The value of M used is that of the repeat unit of the molecule.

3.2.3 Double Refraction A consequence of the existence of more than one index of refraction in a material is the phenomenon of double refraction. When a beam of unpolarized light is directed on to a crystal of certain materials (such as calcite) from an appropriate direction, it is found that there are two refracted beams instead of the usual one. This effect can be seen as a duplication of the image when examining an object through a calcite crystal (Fig. 3.1). These two beams can be shown to be polarized at right angles to one another by viewing the image through a sheet of polaroid; as this is rotated, first one image will be extinguished so that only one image remains (Fig. 3.2) and on rotation through a further 90° the other image will disappear. If the angles of refraction are measured for both beams, it is found that Snell's law of refraction holds for only one of them; this ray is called the ordinary or '0' ray and the other is called the extraordinary or 'e' ray. The ordinary ray lies in the plane of incidence but this is not in general true of the extraordinary ray. If the incident light is normal to the surface, the ordinary ray will pass through without deviation but the extraordinary ray will be refracted at an angle to it such that, on rotation of the crystal, the ordinary ray will remain stationary whilst the extraordinary ray revolves around it. The double refraction effect in uniaxial crystals disappears when the incident light travels parallel to the optic axis.

78 B. P. Saville

FIG. 3.1. Double refraction by a calcite crystal.

The explanation for this behaviour lies in the fact that unpolarized light contains components vibrating in all directions, and within the material the light vibrating in different directions can travel with different velocities through it depending on the refractive index for that particular vibration direction. In a uniaxial material, light vibrating in a plane at right angles to the optic axis encounters the same refractive index for all directions that are at right angles to the optic axis. However, light vibrating in a plane parallel to the optic axis encounters a changing refractive index as the direction of vibration is rotated within this plane. Light vibrating in such a plane will spread through the material with an ellipsoidal wavefront owing to the difference in velocity with direction. This is in contrast to the situation in an isotropic material where the light has a spherical wavefront. In a material that exhibits double refraction the component of the incident light vibrating perpendicular to the optic axis propagates through the material with a spherical wavefront (Fig. 3.3(a)), behaving as if the material were isotropic and


FrG. 3.2. As Fig. 3.1 but viewed through a polaroid sheet.

thus giving rise to the ordinary ray. The component vibrating parallel to the optic axis propagates with an ellipsoidal wavefront (Fig. 3.3(b»; the resultant direction of the ray is at an angle to the direction of the ordinary ray since in such cases the ray direction is no longer perpendicular to the wavefront. In the case of biaxial materials there is in general no ordinary ray for the reason that the light waves propagate with ellipsoidal wave fronts in nearly all directions, so there are usually two extraordinary rays instead.

3.2.4 Dispersion of Refractive Index The refractive index of a dielectric material is not a fixed constant but has a value that varies with the frequency of the incident radiation. For a colourless transparent material the index decreases with an increase in wavelength. In regions of the spectrum away from any absorption bands, the variation of refractive index with wavelength can be described by the Cauchy relation:

80 B. P. Saville

---">..+--'''------~+_'O---''''--f_'''- ........ wave front ./' ! I , I

--............ ra~ ~irection ~

(al ordinary ray (b) extraordinary ray

FIG. 3.3. Wavefronts of the rays in double refraction.

n = A + B/).} + C/).4 (3.3)

where A, Band C are constants characteristic of the substance in question. Each principal refractive index of an anisotropic material also varies in the same way.

The dispersion of the refractive index is a measure of the degree of variation of the refractive index with wavelength. Total dispersion is the term applied to the numerical difference between the refractive indices of the material for wavelengths at the opposite ends of the visible spectrum. Relative dispersion is given by

(j = nF - nc no - 1

(3.4)

where nF, nc and no are the refractive indices for the reference wavelengths F (486·1 nm), C (656·3 nm) and D (589·3 nm).

For most anisotropic materials the variations in the individual refractive indices with wavelength follow very similar curves, so the difference between them, i.e. the birefringence, remains substantially the same throughout the visible spectrum. However, some materials exhibit a significant dispersion of the birefringence in that the curves of the individual indices follow diverging or converging paths. A wedge of such material, when viewed in white light, would show a sequence of colours different from the normal Newton's scale because the path difference at a given point would depend on the wavelength as well as the thickness. Such colours are referred to as anomalous interference colours.


3.3 ELLIPTICALLY AND CIRCULARLY POLARIZED LIGHT

The general description of polarized light that has passed through a thin birefringent plate as 'elliptically polarized' is a convenient way of denoting the fact that it consists of two components vibrating at right angles to one another. This concept and notation finds it greatest use in situations where polarized light traverses more than one thin plate, in which case analysis by resolution of the beam into its individual components becomes quite complex.

When a beam of monochromatic polarized light is incident on a thin birefringent plate it is in general split into two components which are polarized at right angles to one another. These two components are coherent since they are derived from the same incident beam, but they are now out of phase with one another because of the different optical paths traversed. It is possible to regard the resultant of these two components as elliptically polarized light in that its direction of vibration and amplitude vary with time such that its electric vector traces out an ellipse. This is the general case; in special cases the ellipse can reduce to a circle or a straight line.

The two component waves can be represented by the formulae

y a sin 00

z = b sin (00 + 0) (3.5)

where y and z are the electrical displacements at any instant, a and b are the amplitudes, 00 is the common phase angle and 0 is the phase difference. When combined they give

• 2 y2 Z2 2yz sm 0 = - + - - - cos 0

a2 b2 ab (3.6)

which is the equation for an ellipse. If the amplitudes of the two waves are equal, i.e. a = b, this corresponds to the frequently met case where a birefringent specimen is oriented at 45° to the polarizer. The effect of phase difference on the resultant vibration for this particular case is shown in Fig. 3.4. Simpler forms of vibration occur when cos 0 is either 0 or 1. When the two components are in phase the phase difference 0 is either 0 or an integral multiple of 2lT and the equation then reduces to y = z which is that of a straight line, so the light is linearly polarized. If o = IT the equation also reduces to that of a straight line y = z with an opposite sign of slope and a direction altered by 2e. When 0 = IT/2 or 3lT/2

82 B. P. Saville

FIG. 3.4. Variation of elliptically polarized light with phase angle 8.

the equation reduces to that of a circle l + Z2 = a2 and the light is then said to be circularly polarized. The direction of motion of the vector in the ellipse or circle is not given by the ellipse equation; this direction reverses at D = IT because a phase lag of between IT and 2lT can also be thought of as a lead by the other component of between 0 and IT.

Rotation of the resultant clockwise (looking at the source) is conventionally termed right elliptically polarized light, and anticlockwise rotation is termed left elliptically polarized.

The resultant that occurs when linearly polarized light passes through two birefringent plates, which individually would give rise to elliptically polarized light having opposite rotations, is linearly polarized light if the ellipses described by the electric vectors are identical in all other aspects.

3.4 THE UNIAXIAL INDICATRIX

A convenient representation of the optical properties of an anisotropic material is the optical indicatrix. This is a plot in three dimensions of the variations of refractive index and vibration direction of the light passing through the material. With its use it is possible to calculate the refractive index encountered by both the extraordinary ray and the ordinary ray in a uniaxial material for unpolarized light incident from any direction.


For uniaxial materials the indicatrix is an ellipsoid of revolution with the radius of the circular section directly proportional to the ordinary refractive index and the length of the axis of revolution directly proportional to the extraordinary refractive index. The refractive index encountered by polarized light vibrating in a particular direction is given by the length of the radius vector of the indicatrix in that direction. For instance, in Fig. 3.5 the refractive index for light vibrating in the direction A-A' is ne and for light vibrating in the direction B-B' it is no as it is for any light vibrating perpendicular to the optic axis. For any other vibration direction the refractive index is intermediate between the two principal values ne and no.

In order to determine the effect that the material has on a parallel beam of light travelling through it in a general direction, it is necessary to take a central section of the indicatrix perpendicular to the wave normal of the light. This is the section of the indicatrix containing the origin which would be cut by the plane of the wave fronts in the crystal. The two axes of this elliptical section define the permitted vibration directions for the wave normal direction whilst the lengths of the respective semi-axes are proportional to the appropriate refractive indices. In the case of the uniaxial indicatrix, one axis of the elliptical section must always lie in the circular section with radius no; hence, regardless of the direction in which light traverses a uniaxial material, one of the two components that it is resolved into will always encounter a refractive index no. In the special case where the wave normal coincides with the optic axis, the elliptical section reduces to a circle of radius no; so there is no difference in the velocity oflight with vibration direction and the material behaves as if it were an isotropic material with a single refractive index no.

3.5 THE BIAXIAL INDICATRIX

Biaxial materials are characterized by three principal refractive indices, na , np and nr in order of increasing magnitude, and an appropriate indicatrix can be·constructed, in a similar manner to that for uniaxial materials, with semi-axes proportional to these refractive indices as shown in Fig. 3.6.

The figure is now no longer rotationally symmetric about any of the principal axes, so none of these is an optic axis as is the case for uniaxial materials. As np is the intermediate principal refractive index, and as the

84 B. P. Saville

optic axis

!

plan elevation

FIG. 3.5. Uniaxial indicatrix for optically negative material; ne < no.

index in the a-r plane varies continuously between the highest and the lowest values of the refractive index, there must be a radius of that ellipse equal to nfl' This means that there is a plane through the indicatrix of circular cross-section with radius nfl. The direction perpendicular to this section is an optic axis because light vibrating in the section perpendicular to this direction encounters the same refractive index in all vibration directions. There are two optic axes that fill this requirement, symmetrically disposed about the r vibration direction in the a-r vibration plane, hence the term 'biaxial'.

optic

elevation

FIG. 3.6. Biaxial indicatrix.


3.6 METHODS OF PRODUCING POLARIZED LIGHT

An optical device that produces polarized light from an input of unpolarized or natural light is termed a polarizer. Practical devices for producing polarized light make use of one of three different physical mechanisms, all relying on some form of asymmetry in the process. The three processes are selective absorption (dichroism), reflection and birefringence. Some polarization is also produced when light is scattered, but this is not usually employed as a way of producing polarized light.

3.6.1 Polarization by Reflection Light reflected from polished flat surfaces is partially linearly polarized. The state of polarization of the reflected light is dependent on the angle of incidence of the light and on the refractive index of the reflecting surface (see Fig. 3.7).

The reflected light is polarized to the greatest degree when the angle between the reflected and refracted rays is 90°. The angle of incidence (0) that gives this, and hence the maximum polarization, is known as the Brewster angle, and this is related to the refractive index of the refracting medium:

tan 0 = n (3.7)

where n is the refractive index of the reflecting medium and the light is travelling from air to the medium. The direction of vibration of the

air

FIG. 3.7. Polarization by reflection.

86 B. P. Saville

polarized light produced is parallel to the reflecting surface; this fact is useful for determining the vibration direction of an unknown polarizer by viewing a source of reflected light through it whilst the polarizer is being rotated. A minimum intensity will be observed when the vibration direction of the unknown polarizer is at right angles to the reflecting surface.

The problem of using this phenomenon is that of constructing a practical polarizer since the reflected beam is weak although highly polarized whereas the refracted beam although strong is only partially polarized. However, polarizers have been constructed on this principle by passing the light through a succession of glass plates arranged at the Brewster angle.

3.6.2 Polarization by Birefringence Polarization by birefringence makes use of the fact that unpolarized light entering a doubly refracting crystal is resolved in two linearly polarized components whose vibration directions are mutually perpendicular. Practical devices rely on isolating one of the two components by making use of the fact that the refractive index of the crystal is not the same for the two components. The most well known polarizer of this type is the Nicol prism which was the usual form of polarizer found in scientific instruments before the advent of Polaroid sheet. The Nicol prism is made from a calcite crystal which is split diagonally and cemented back together with Canada balsam. The paths of the two rays are arranged so that one of them is totally internally reflected at the Canada balsam/calcite interface, so leaving the other linearly polarized ray to pass through the prism.

3.6.3 Polarization by Selective Absorption Some anisotropic materials show different absorptions for the '0' and 'e' rays. This effect, known as dichroism, is very marked in some materials, such as the naturally occurring mineral tourmaline, and it provides a simple way of producing almost completely linearly polarized light by removing one of the rays by absorption. Synthetic materials of this type produced by the Polaroid Corporation are now almost universally used in optical instruments employing polarized light in the visible spectrum. These materials are produced by orientating the molecules in a polymer sheet by stretching and then rendering these dichroic by the use of suitable dyes which align themselves with the polymer chains.


3.7 TYPES OF BIREFRINGENCE

In polymeric materials there are a number of different ways in which birefringence can arise; these will be described under the various headings.

3.7.1 Orientation Birefringence Orientation birefringence is produced by the alignment of molecules that are themselves optically anisotropic, as is generally the case for polymer chains. Polymer chains can be considered to have a different refractive index parallel to the chain from that perpendicular to it. When the chains are randomly arranged, as in a melt or in the interior of mouldings, the net refractive index is intermediate between the two extremes and is the same for all directions in the material. When the randomness is disturbed by alignment of the chains, the refractive index of the material can vary with the direction in the material. Molecular alignment can occur in both crystalline and amorphous polymers, either as a result of a deliberate drawing process, as in the manufacture of fibres and films, or as a by-product of the deformation that occurs in many processes such as extrusion or injection moulding. Because of this connection between orientation and birefringence, attempts are often made to measure birefringence as a means of quantifying orientation. However, the existence of other sources of birefringence as noted below can affect the interpretation of such measurements.

3.7.2 Strain Birefringence Strain birefringence is found in materials that are under stress. The superimposed stress can alter the distance between the atoms of the material, thus changing the polarizability of the bonds in the direction of the applied stress, and hence creating a difference in refractive index in the material between the stress direction and directions perpendicular to it. This effect can occur equally well with molecules that are optically isotropic in the unstrained state and those that are naturally anisotropic. The effect can be seen, for instance, in glass subject to stress. In practice, with polymers it is difficult to distinguish between orientation and strain birefringence, because applied forces can both align molecules that are in a random configuration and deform any molecules that are already aligned in the direction of the force. Thus, when all the polymer chains in a sample are parallel to one another, any extra strain in that direction can only increase the atomic separation in the chains.

88 B. P. Saville

3.7.3 Form Birefringence Form birefringence is a phenomenon that is found in materials having two or more separate phases with different refractive indices. If one phase is in the shape of rods or plates with their smallest dimension less than the wavelength oflight, the refractive index of the whole material parallel to the rods is different from that perpendicular to the rods, even when both phases are themselves isotropic. Effects of this type are found in styrene-butadiene block copolymers? It is possible in some cases to estimate the contribution of form effects to total birefringence by selectively swelling one of the phases in various liquids of different refractive indices; the form birefringence will in theory fall to zero when the refractive index difference between the phases is reduced to zero.

3.8 THE PASSAGE OF POLARIZED LIGHT THROUGH THIN BIREFRINGENT PLATES

When a parallel beam of linearly polarized light is incident on the surface of a thin sample of an anisotropic material it can encounter two different refractive indices in the material depending on the vibration direction of the light (Fig. 3.8(a)). The directions in the material which are associated with these two indices are at right angles to one another. The values of the two refractive indices are determined by the orientation of the indicatrix of the material with respect to the incident light direction.

If the vibration direction of the incident light is parallel to either of these two principal refractive index directions of the sample, only linearly polarized light vibrating in that direction is transmitted. In the more general case where the vibration direction of the light is at an angle to the principal directions, the incident beam is resolved into two separate components vibrating in these directions. The relative amplitude of these two components varies with the angle between the vibration direction of the incident beam and the principal directions in the sample as shown in Fig. 3.8(b). The vibration direction of the incident beam is determined by the vibration direction of the polarizer. Because these two components experience different refractive indices they travel through the specimen at different velocities, the component vibrating along the direction of highest refractive index travelling the slowest. For this reason the direction of highest refractive index in the material is often called the slow direction and the direction of lowest refractive index the fast direction.

Polarized Light: Theory and Measurements

incident

...- polarized light

~ __________ <f.--Sample

elevation

/

" /

/ /

/

"'""~ A sine

(a)

principal

refractive Indices

(b)

incident beam amplitude A

/

n,

A cose

plan

AoO ... ~ /;' analyser vibration

/ direction

(e)

89

FIG. 3.8. (a) Relation between incident polarized light and principal refractive indices of a material. (b) Resolution of incident beam into components by a birefringent material. (c) Analyser passing components (shown for crossed

polars).

90 B. P. Saville

When the two waves are produced on entering the specimen they are in phase, since they are components of the same original vibration of wavelength A; however, the different optical paths that they travel cause them to be out of phase on leaving the specimen. If the thickness of the specimen is t and the two refractive indices are n I and n2 then the optical paths of the two waves are nIt and n2t and the optical path difference between them is (n I - n 2)t. The corresponding angular phase difference is

(3.8)

In order to render this phase difference visible it is necessary to resolve each of the two waves into components whose vibrations are in the same direction. These components are then able to interfere either constructively or destructively with one another depending on the phase difference between them.

To do this we insert another polar, the analyser, after the specimen, but with its vibration direction at right angles to the vibration direction of the polarizer. The analyser passes only that component of the transmitted light vibrations parallel to its permitted vibration direction. A sample viewed in this way is said to be examined between crossed polars.

What is seen when a specimen is examined between crossed polars depends strongly both on the orientation of the specimen with respect to the vibration directions of the polars and on the phase difference introduced into the two waves by the specimen. The effect of rotation of the specimen is to vary e (Fig. 3.8(b )). If e is either 0° or 90° the incident polarized light is not split and all of it will traverse either the fast or the slow direction of the specimen. However, the light is all blocked by the analyser which is at 90° to the polarizer. For values of e between 0° and 90° there is a resolved component parallel to the analyser (Fig. 3.8(c)) equal in both cases to A cos e sin e. This has its maximum value of O' 5 when e is 45°. The practical effect of this is seen when a birefringent specimen is rotated between crossed polars when there is a cyclical change in brightness every 90°. When the specimen is parallel to either the polarizer or the analyser there is complete darkness, termed extinction, and when it is at 45° to either there is a maximum in the brightness. It is in this position that measurement of birefringence is usually made.

The phase difference produced by a specimen is dependent on both


the thickness of the specimen and its birefringence; it can be assessed either qualitatively from the interference colour produced or quantitatively by using a compensator.

It we consider light of only one wavelength (monochromatic), the effect of the phase difference between the two components introduced by the specimen is to produce either constructive or destructive interference between the components. This can only occur when the vibration directions of the waves are parallel to one another, which is the case for the resolved components which pass the analyser. Destructive interference occurs for every whole wavelength path difference between the two components. This means that any specimen having an optical path difference of nA, where n is a whole number, will appear black between crossed polars in monochromatic light. This is reasonable because a shift of a whole wavelength in phase between two waves is indistinguishable from the starting condition. If a specimen that did not produce any path difference, e.g. an isotropic one, is viewed between crossed polars it will appear black.

Changes in path difference can be caused by changes both in birefringence and in thickness. A wedge of birefringent material shows evenly spaced dark bands with increasing thickness where the condition

(3.9)

is fulfilled for different values oft. It is important to distinguish between the two possible factors dictating the observed optical path difference, as this is the quantity that is rendered visible and can be measured between crossed polars.

3.9 POLARIZATION COLOURS

It has been shown previously that, for a birefringent sample viewed in monochromatic light between crossed polars, there is destructive interference when the optical path difference is a whole number of wavelengths. Hence a wedge of such material will show a series of regularly spaced dark bands along it, corresponding to the points where the product of the thickness and the birefringence is a whole number of wavelengths, as shown in Fig. 3.9. However, white light, which is more usually used for examining specimens, contains a continuous spread of wavelengths between about 400 nm and 750 nm, so destructive inter-

92 B. P. Saville

FIG. 3.9. A wedge of oriented Perspex viewed between crossed polars in monochromatic light.

ference will occur at a different path difference for each part of the spectrum. Therefore, at no point can there be total darkness, for a specimen viewed in white light. For a specimen with a given (non-zero) optical path difference there will be subtraction from the incident white light of that wavelength for which the path difference is an integral multiple. Other wavelengths will also be reduced in intensity to a decreasing extent the further they are removed from this particular wavelength. With thicker or more birefringent specimens the path difference can be a multiple of more than one wavelength at the same time. The resulting complex subtraction of wavelengths from the incident white light produces a characteristic colour which can be related by use of a chart to the optical path difference producing it. This identification is only possible at the lower end of the path difference scale.

The sequence of colours produced between crossed polars in white light by a specimen with steadily increasing optical path difference, such as a wedge of birefringent material, is known as Newton's scale of


interference colours. As the path difference increases from zero the scale goes through a colour sequence which ends in a characteristic magenta shade at a path difference that is one wavelength for the middle of the visible spectrum. Mterthis there is a different sequence of colours which again ends in a magenta shade. This sequence is subsequently repeated, with the tints becoming paler at each repetition, until after about five sequences the colours become almost indistinguishable from white light. The repetition within the colour scale and the appearance of the magenta tints allows the division of Newton's scale into orders; a colour appearing between zero path difference and the first magenta tint would be said to be a first-order colour. The white light appearing after the fifth order is known as high-order white and in certain cases it may need to be distinguished from the white that appears at the beginning of the first-order. This can be done by viewing the material between parallel polars or by inserting an additional small fixed retardation plate. In both cases the high-order whites are unaffected by the change whereas the first-order white changes considerably.

The actual interference colours observed in a material depend on the spectral content of the white light used, and on whether the birefringence of the material is independent of wavelength. A strong body colour can also affect interference colours. Any colour that does not match Newton's scale for any reason is referred to as an anomalous colour. In order to determine the order of an interference colour and its place in the colour sequence, use is made of a coloured chart showing the first few orders, known as the Michel-Levy chart.

When a material is viewed between parallel polars there is an interchange of conditions for constructive and destructive interference compared with the case for crossed polars, so the colour sequence with increasing path differences is altered. The colour at a given path difference is then complementary to that observed between crossed polars.

3.l0 RELATION BETWEEN ORIENTATION AND BIREFRINGENCE

In many practical applications, such as film and fibre manufacture, birefringence is used as a measure of molecular orientation in a product. To specify completely the orientation of the polymer chains

94 B. P Saville

produced by a drawing process would require distribution functions that accounted for the orientation of every one of the molecules in the sample. It is not possible to obtain all of this information from a single measurement such as birefringence. In most cases use is made of orientation factors, such as that of Hermans. These relate a fixed direction in the material, such as the draw direction, to the chain direction of oriented molecules or to a particular crystallographic direction of interest.

For a drawn fibre where the molecules are considered to be aligned with the draw direction but only randomly arranged in the cross section, the optical properties of the system can be specified by only two refractive indices, nl parallel to the fibre and n2 perpendicular to the fibre. The Hermans orientation factor fH is then related to the birefringence by the relation

n l - n2

~o (3.10)

where ~o is the intrinsic maximum birefringence, i.e. the maximum possible value of (n I - n2) which corresponds to the case where all the molecules are perfectly aligned. The factor is related to orientation by

'H -_ 3 cos2 ¢ - 1 JI 2 (3.11)

where cos2 ¢ is an average value for all the polymer chains, being the angle between the polymer chain and the fibre axis. It can also be considered to be equivalent to a notional fibre that has all its chains oriented at this angle to the fibre axis. If all the polymer chains were aligned parallel to the fibre axisfH would be 1. For an isotropic system where there is no orientation fH would be 0, and if the chains were aligned perpendicular to the fibre axis fH would be -!.

In the case of an oriented polymer film which can be drawn to different extents in the two directions, it may be necessary to consider three refractive indices in order to fully characterize the orienta tion. The individual molecules can be considered to have only two refractive indices, one parallel and one perpendicular to the chain; these are associated with the polarizability along the chain and the average polarizability across it. If, however, the polymer chains have a tendency to be grouped into sheets (by hydrogen-bonding for instance), all of the three refractive indices of the polymer crystal may have to be considered.


The biaxial nature of a drawn film arises from the different orientation of the chains in the three different planes rather than from the polymer having an optically biaxial crystal. In the case of a strictly uniaxially drawn film (see Fig. 3.10), two of the refractive indices will be the same, that in the transverse (n2) and that in the direction perpendicular to the sheet (n3). The birefringence in the y-z plane (nl - n2) is then all that is needed to specify the orientation. White and Spruie1l3 have developed an orientation factor for this case given by

(3.12)

where <P is the angle between a reference direction and the chain. In a biaxially drawn film the orientation tends to be planar, i.e. the chains are all oriented perpendicular to the x-axis but the orientation in the y-z plane can be random depending on the balance between the draw ratio in the machine direction and that in the transverse direction. The disposition of the chains in the uniaxial and biaxial cases is shown diagrammatically in Fig. 3.11. In such a case the birefringence of the y-z plane can only give information about the balance of the orientation between the machine and transverse directions. A balanced film with equal draw in both of these directions would show no birefringence in this plane. To characterize the film the birefringence has to be measured for both draw directions with respect to the direction x perpendicular to the plane of the film, i.e. (nl - n3) and (n2 - n3). White and Spruie1l3

have proposed the use of two orientation factors in this case:

(3.13) 1;, = 2 cos2 <Ply + cos2 <Plz - 1

where <Ply and <Plz are the angles between the polymer chain and the y-

FIG. 3.10. Relationship between refractive indices and draw direction for a biaxially oriented material.

96

n ,

n,

B. P. Saville

n,

n, uniaxially

drawn material n3 ~ 0 0 0 c:::::J 01 1000

biaxially

drawn material

FIG. 3.11. Schematic diagram showing orientation of the polymer chains in a drawn material.

axis and the z-axis respectively. For uniaxial orientation in the machine (z) direction/, = 1,.1;, = O. For uniaxial orientation in the transverse (y) direction/, = 0,.1;, = 1. For a film with equal biaxial orientation

/, = I' = 1 - 3 cos2 ¢lx z Jy Z (3.14)

where ¢lx is the angle between the chains and the x-axis. If the equal biaxial orientation is planar, i.e. ¢lx = 90°, then j,.=.I;, =!. The orientation factors are related to the measured birefringences by the equations

fz nl - n3

.6.0 (3.15)

.1;,= n2 - n3

.6.0

The main difficulty which arises when using orientation factors is that of obtaining a reliable value for .6.0' the maximum birefringence.

3.11 THE POLARIZING MICROSCOPE

In principle any light microscope could be converted for use as a polarizing microscope by equipping it with two pieces of polaroid sheet, one placed before and one after the specimen. However, in order to interpret the image formed in these circumstances it is necessary to


perform certain instrumental tests such as rotation of the specimen or insertion of fixed retardation plates. Therefore instruments intended for routine observations using polarized light have a number of extra features above those found on microscopes used for common light only, and for measurements using polarized light further accessories are needed. The following are some of the commonly encountered accessories.

Removable Polarizer and Analyser The provision of polars that can be easily inserted or removed allows the microscope to be changed rapidly from a polarizing instrument into one for common light observation. The analyser is usually mounted on a slide with two positions: analyser in and analyser out. The polarizer alone is used for viewing specimens in polarized light, such as when measuring the separate refractive indices of a birefringent material.

Rotating Stage Rotation of the specimen is needed to determine the difference between a specimen that appears black between crossed polars owing to its being isotropic and one that appears black owing to its being in an extinction position. It is also used to find the extinction direction of a specimen, and hence the position of maximum brightness which is at 45° to this. The stage is usually equipped with an angular vernier scale for this purpose, but the more expensive microscopes have a stage with an indent which can be engaged at the extinction position and which then also engages at every 45° rotation increment from this position.

The use of a rotating stage means that a method has to be provided for centring the axis of rota tion of the stage with respect to the optical axis of each objective in use. Often this is done by having a centring mechanism on the stage itself, but on more expensive microscopes it is preferred to give the stage very accurate bearings and to make provision for adjusting each objective lens separately. This adjustment is sometimes provided for by mounting the objectives on separate individual carriers with their own centring screws rather than on a revolving nosepiece.

Body Slot The body slot is situated between the objective and the analyser and is made to standard dimensions to permit the insertion of compensators and fixed retardation plates. The orientation of the slot is at 45° to the

98 B. P. Saville

vibration directions of the polarizer and analyser so that a plate inserted into it is either parallel or perpendicular to a specimen set at its maximum brightness position.

Strain Free Optics Any optical component situated between the polarizer and analyser which has strain in it will show a certain amount of brightness between crossed polars due to the strain birefringence in the glass, so reducing the degree of extinction when the polars are crossed. Therefore strainfree optical components need to be specially selected for a polarizing microscope. Such objectives should not be used with microscope hotstages.

Bertrand Lens The Bertrand lens is an auxiliary lens situated above the analyser which allows the back focal plane of the objective to be viewed. It is used for the conoscopic examination of materials, a technique that is much used in optical mineralogy.

Eyepiece Crosswires The crosswires in the eyepiece are arranged to be parallel to the vibration directions of both the analyser and the polarizer, so indicating their orientation relative to the specimen. An eyepiece equipped with crosswires has therefore a projection which fits into a special slot in its tube in order to align it correctly.

Rotating Analyser A rotating analyser fitted with an accurate angular scale is necessary when using the Senarmont method of compensation for measuring small path differences. In this case it is used in conjunction with a special quarter-wave plate. It is also necessary to be able to rotate at least one of the polars to be able to obtain precise extinction.

Monochromatic Light Monochromatic light is usually achieved with an appropriate filter used in conjuction with the white light from a tungsten lamp. Monochromatic light of greater intensity can be obtained by using a mercury discharge lamp together with a filter for one of the peaks in the spectrum of the light such as green 546 nm. Its use is essential for accurate measurements of optical path differences.


Powerful Light Source The use of crossed polars, particularly together with monochromatic filters, considerably reduces the intensity of an image. Therefore a powerful light source is useful in polarized light work for examining specimens of weak birefringence and especially for photomicrography.

3.12 MEASUREMENT OF OPTICAL PATH DIFFERENCE

The first step in measuring optical path difference in a polymer is to mount a suitably thin specimen in a liquid of matching refractive index. Fibres and films can usually be mounted whole, but most mouldings or extrusions will require sectioning first. Unfortunately the sectioning process introduces strain into a specimen which appears as a modification to its birefringence, so interfering with the measurements being made. One possible way round this problem is to cut sections from the specimen of different known thicknesses when the induced component, which affects the outer layers only, will remain constant so that the true birefringence can be obtained from the slope of a plot of optical path difference versus thickness. In other cases it may be possible to produce a strain-free section by either grinding and polishing the specimen to a thin section or polishing both surfaces of a thick section. The polishing process produces lower levels of strain which have a more random distribution in the specimen than is the case in sectioning.

The actual measurement on the mounted specimen is normally made between crossed polars, with the specimen at the position of maximum brightness. As it is difficult to determine this position with any accuracy, the extinction position is first located and the specimen is then rotated 45° from this position. A second set of measurements is usually made with the specimen at one of the other positions of maximum brightness which are 90° or 180° to this. The direction of the extinction position relative to a chosen direction in the specimen such as the machine direction gives the preferred orientation of the molecules within the specimen, and this itself can be a measurement of interest. For instance, the orientation direction and machine direction in a drawn film do not necessarily coincide, and the angle between them may vary across the width of the film.

Measurement of optical path difference in the microscope is commonly carried out with some form of compensator which is introduced into the light path between the polars, usually by way of the

100 B. P. Saville

body slot. The orientation of the compensator with respect to the specimen is arranged so that the slow direction of one is parallel to the fast direction of the other. By this means the variable birefringence of the compensator can subtract from that of the specimen and thus produce zero path difference by adjustment.

For accurate work the measurement should be made in monochromatic light, but for path differences of more than one wavelength it is only possible to distinguish the zero-order fringe by using white light. The reason for this is that in monochromatic light there is a dark fringe for each and every whole number of wavelength path differences, so if the path difference of the specimen is 1·25). then there is extinction for compensator settings ofO· 25)" 1·25).,2·25).,3·25)' etc. Therefore for path differences greater than one wavelength it is necessary to determine the whole number of wavelengths involved as well as the fractional part.

Many compensators produce only a narrow band of compensation which does not fill the field of view, so it is necessary in these cases to centre the crosswires in the eyepiece on a part of the specimen that is of interest and to make the measurement only at that point. It is often helpful in these cases to reduce the field of view by closing down the field iris.

3.13 COMPENSATORS

3.13.1 The Quartz Wedge The quartz wedge is the simplest form of compensator; the necessary variation in optical path difference is produced by the change in thickness along the wedge. For quantitative work the wedge has a scale directly engraved on it, graduated either in path difference or in arbitrary units, in which case the scale has to be calibrated before use. In order to read the scale on the wedge, some special arrangement has to be used to bring the plane containing the wedge into focus. This is usually done with a slotted eyepiece known as a Wright eyepiece.

To use the wedge, the specimen is placed on the stage of the microscope between crossed polars and rotated 45° from extinction to its position of maximum brightness. The quartz wedge is then inserted so that it is at45° to the vibration direction of the polars, and the change of colour produced by the combination of specimen and wedge is noted. If the colours ascend Newton's scale from the original colour of the specimen, the path difference of the specimen and wedge are adding


and the specimen must be rotated through 90° . In this position the colour produced should descend the scale until extinction is produced, which is the point of compensation.

For calibration of the wedge, it is viewed on its own between polarizer and analyser in monochromatic light, when extinction bands spaced exactly one wavelength apart will be seen. Between crossed polars these will correspond to path differences of lit, 2A etc., and between parallel polars Al2, 3M2 etc. The optical path difference can then be plotted against the scale reading, the value of A being given by the wavelength of the monochromatic light used.

The range ofthe quartz wedge can be increased by inserting a plate of fixed path difference into the body slot, with its slow direction parallel to that of the wedge. This adds a constant path difference to the wedge; the value of the fixed plate can first be determined using the wedge. For measuring small values of path difference a first-order plate can be inserted in the body slot with its slow direction at right angles to that of the wedge, so subtracting a fixed path difference. This then gives a zeroorder band not previously available since the thin edge of the wedge is always missing owing to grinding difficulties.

3.13.2 The Babinet Compensator The Babinet compensator consists of two quartz wedges of equal angle positioned one above the other as shown in Fig. 3.12. One of the wedges is capable of being traversed relative to the other by a micrometer screw. One wedge is cut with its long dimension parallel to the optic axis of the quartz, and the other has the optic axis of the quartz aligned across the wedge. In the figure, lines indicate where the optic axis of the quartz is parallel to the paper, and dots show where it is perpendicular. This arrangement means that the fast component of one wedge opposes the slow component of the other. Where the thickness of the two wedges is the same, as will occur in the centre when the movable wedge is at zero, there is no resulting optical path difference at that point. When the compensator is positioned in the 45° position between crossed polars, a series of equally spaced extinction fringes will appear across the width

FIG. 3.12. Babinet compensator.

102 B. P. Saville

of the compensator at the points where the optical path difference is a whole number of wavelengths. In white light these fringes will be coloured, with the exception of the central fringe, where the path difference is zero. In monochromatic light all the fringes will appear identical, so there is then no way of distinguishing which is the zero fringe.

In use, the presence of a birefringent sample in the 45° position will cause the central zero fringe to be shifted along the compensator, the direction of the shift depending on whether the slow direction of the sample is parallel or perpendicular to the compensator. The displacement of the fringe and hence the path difference is determined by bringing the zero fringe back to the centre of the field of view with the micrometer adjustment. This adjustment is calibrated before use in monochromatic light in a similar way to the calibration of the quartz wedge.

3.13.3 The Berek Compensator The Berek compensator consists of a thin calcite plate cut normal to the optic axis and mounted on a spindle so that it can be tilted on either side of the horizontal position up to an angle of about 20°. Rotation of the plate about the horizontal axis provides an increase in optical path difference, partly through increased thickness and partly through a change in angle of its indicatrix giving an increased birefringence. The angle of rotation of the plates can be accurately measured on an external scale. This scale can be calibrated for optical path difference using monochromatic light, in a manner similar to that for the quartz wedge, but the relationship between optical path difference and angle will not be linear. However, the theoretical form that this curve should take is known. Because of this, these instruments are usually supplied with tables listing the optical path differences for different values of rotation angle. The range of this compensator is usually up to three or five orders. Because of the large dispersion of birefringence in calcite, versions of this compensator are also made from other minerals such as magnesium fluoride.

3.13.4 The Ehringhaus Compensator The Ehringhaus compensator is somewhat similar in principle to the Berek compensator but with the difference that two crystal plates of either calcite or quartz are used instead of one. These are both cut parallel to the optic axis of the material and cemented together with


their axes at right angles to one another so that the resultant optical path difference is zero when the plate is horizontal. Quartz plates are used for path differences of up to 41\., and compensators using calcite plates are capable of measuring path differences of up to 1221\..

3.13.5 The Brace-Kohler or Elliptic Compensator This is a form of rotary compensator in which a thin mica plate of accurately known path difference is rotated about the vertical axis of the microscope in order to achieve extinction. It is usually available for three different maximum measuring ranges (/tI30, /tI20, /til 0) and is the most sensitive form of compensator for very low path differences.

When carrying out measurements the compensator, positioned in the body slot, is first set to its extinction position and then the specimen is inserted in one of its 45° positions. The compensator is rotated until extinction is again obtained. The angle through which the compensator has been rotated is then related to optical path difference (OPD) by the expression

OPD = C sin 2(} (3.16)

where () is the angle through which the compensator has been rotated from its extinction position and C is a constant for the particular compensator. Maximum compensation is obtained when the mica plate has been rotated through 45°. In use, after extinction has been obtained for one position of the specimen, the specimen is rotated through 90° and a further reading of the compensator is made which should be equal and opposite to the first. The difference between the two readings is then 2(}. Following this procedure eliminates zero errors.

3.13.6 Senarmont Method The Senarmont method uses a fixed quarter-wave plate in the tube slot and a rotating graduated analyser. The quarter-wave plate is a special one of high accuracy which has its slow vibration direction parallel to that of the polar; alternatively a rotatable quarter-wave plate may be used, set to give extinction between crossed polars. For accurate measurement the plate must be used with monochromatic light of the wavelength for which it is graduated.

In use, the analyser is set to give extinction at its zero position and the specimen is rotated to one of its 45 ° positions. The quarter-wave plate is then inserted in the body slot and the analyser is rotated so as to bring the centre of the specimen back to extinction. The nearest extinction

104 B. P. Saville

position to the zero setting is chosen. The specimen is then rotated through 90° and the analyser returned to zero, and then the other extinction position is found by rotating the analyser in the opposite direction to that for the previous reading. The two readings should be of nearly equal magnitude and of opposite sign; the sum of the two angles gives the angular phase difference produced by the specimen directly. If the angle of rotation of the analyser for compensation is (), the phase difference is 2() and the optical path difference is given by

OPD = A 2() (3.17) 360

where A is the wavelength of the monochromatic light.

3.14 DISPERSION OF BIREFRINGENCE

A problem which can be encountered with birefringent polymer samples of high optical path difference, of the order of several wavelengths, is that the variation of the birefringence of the compensator with the wavelength of the incident light may not be the same as that of the specimen. This means that, although the optical path difference of the sample may be matched by that of the compensator for one wavelength, it will not necessarily be matched at other wavelengths. Therefore the interference colours produced by the combination of the path difference of the specimen and that of the compensator will appear anomalous. As the total path length in the two materials increases, the discrepancy in path differences across the spectrum also increases, so in white light the black zero-band becomes increasingly coloured and thus indistinguishable from the adjacent bands delineating the different orders. With further increases in path differences, the band adjacent to the true zero-band becomes black; this is sometimes termed fringe jumping. It has been calculated for drawn polyethylene terephthalate paired with a quartz compensator that the black fringe moves up one position from the true zero about every six orders.4

Unfortunately, in order to measure the path difference by compensator it is necessary to identify the zero-order position where the optical path difference of the compensator just matches that produced by the specimen. There are two possible ways of overcoming this problem:

(1) Construct the compensator from the same material as that of


which the birefringence is being measured. This could be a fixed value compensator of a known but lower path difference than that of the specimen, used in conjunction with a standard variable compensator. This would reduce the problem to that of measuring a small path difference where dispersion does not render the zero fringe unrecognizable.

(2) With fibres and films it is possible, using a sharp blade, to cut the end of the fibre or the edge of the film into a wedge so that the thickness tapers to zero. If the wedge is then observed while the compensation is increased, the zero-order fringe can be observed to start at the thin end of the wedge where it is truly black, and it can be followed up the wedge until it reaches the full thickness of the specimen. This allows the true zero-band to be identified despite the colour changes that it undergoes with increasing total path difference.

Alternatively, a non-microscopical method of determining birefringence which does not use a compensator can be used, particularly with fibres and films.

3.15 SPECTROPHOTOMETRIC METHOD

The spectrophotometric method is a means of measuring the optical path difference of a film or fibre by using a spectrophotometer operating in the visible region of the spectrum. The ability to do this depends on the fact that, although materials of higher optical path difference appear white ('high order' white) when viewed between crossed polars, the spectrum contains a number of discrete extinction bands. These are spread throughout the visible region of the spectrum, so they do not noticeably affect the overall colour. The discrete extinction occurs when the optical path difference is an integer multiple of a wavelength in the visible region.

If a spectrum is recorded from such a material between crossed polars, it will show several absorption peaks all fulfilling the condition

nA = OPO (3.18)

where n is an integer and A is the wavelength of the peak. If it is assumed that the birefringence does not change between adjacent peaks (i.e. negligible dispersion over the small range of A) then

(3.19)

106 B. P Saville


where Al and A2 are the wavelengths of adjacent peaks. This can be rearranged to give

A2 n = ---Al -A2

(3.20)

from which the optical path difference can be calculated. This should be done at long wavelengths since the effects of dispersion are less marked there.

3.16 THE WEDGE METHOD

A method of optical path difference measurement which is applicable to transparent specimens, particularly those of low birefringence, is to cut a large wedge of the material. This can be up to 10 mm deep at the thick end, depending on the transparency of the material involved and the working distance of the objective to be used. When examined between crossed polars the wedge will show a fringe for each whole wavelength increase in optical path difference, as shown in Fig. 3.13. The linear distance between the fringes are measured, preferably in monochromatic light, and the vertical distance between two adjacent fringes can then be worked out if the wedge angle is known (Fig. 3.14). From the thickness of material that produces a path difference of one wavelength the birefringence of the material can be obtained. This method avoids the need to cut a section and so introduce strain. It is particularly useful for transparent thermosetting resins such as polyesters. The cut edges of the specimen must be polished by the techniques that are used to produce specimens for reflected light microscopy, and the whole specimen can be immersed in a suitable liquid for measurement, to avoid refraction effects.

8

FIG. 3.14. Height difference between fringes in the wedge method.

108 B. P. Saville

FIG. 3.15. Principle of the use of the Abbe refractometer on birefringent films; ec is the critical angle, n' is the refractive

index of the prism.

film sample

\

n'

3.17 USE OF THE ABBE REFRACTOMETER

n

Use of the Abbe refractometer for measuring the three principal refractive indices of a drawn films relies on the fact that to determine refractive index the refractometer measures the critical angle for a transition from a material oflowerindex n to one ofa higherindexn'. At the critical angle light is travelling parallel to the prism surface since that is the maximum angle at which light can be refracted in the prism (Fig. 3.15). Therefore, if a film is inserted into the refractometer, light will travel almost parallel to its surface and so will encounter the refractive index perpendicular to the plane of the film (nx ), as well as either ny or nz depending on the orientation of the film to the light paths. The two refractive indices will give rise to rays with slightly different critical angles and which are polarized at right angles to one another, so a rotatable polar placed after the prism can be used to distinguish between the two and thus measure their values directly. The third refractive index can be obtained by rotating the film through 90° and repeating the measurement.

REFERENCES

1. Hamza, A. A. & Sikorski, 1., Optical anisotropy of poly(p-phenylene terephthalamide) fibre. J Microscopy, 113 (1978) 15.

2. Folkes, M. 1. & Keller, A., The birefringence and mechanical properties of a 'single crystal' from a three block copolymer. Polymer, 12 (1971) 222.


3. White, J. L. & Spruiell, J. E., The specification of orientation and its development in polymer processing. Polymer. Eng. and Sci., 23 (1983) 247.

4. Hartshorne, N. H., Modem applications of polarization microscopy. Science Progress, 50 (1962) 11.

5. Samuels, R. J., New techniques for the characterization of orientation in semicrystalline polymer moulding. Polym. Eng. and Sci., 23 (1983) 257.

BIBLIOGRAPHY

Hartshorne, N. H. & Stuart, A., Crystals and the Polarizing Microscope, 4th edn. Elsevier, New York, 1970. Wahlstrom, E., Optical Crystallography, 5th edn. Wiley, New York, 1974. Born, M. & Wolf, E. Principles o/Optics, 4th edn. Pergamon, London, 1970.

4

Polarized Light: Qualitative Microscopy

B. P . SAVILLE

Institute of Polymer Technology, University of Technology, Loughborough, UK

4.1 INTRODUCTION

This chapter is concerned with the uses that are made of polarized light in the study of polymer structure. These uses are determined by the type of polymer under examination, i.e. whether it is semicrystalline or amorphous. Semicrystalline polymers crystallize from the melt, in the absence of significant shear or elongational flow, in the fonn of a characteristic structural entity called a spherulite. Variation in size, shape and type of spherulite with temperature and flow conditions during crystallization is a powerful diagnostic indicator ofthe change in the factors throughout a complex moulded product. Hence the study of spherulitic texture in these materials will reveal the structural changes caused by varying production conditions. In fact the study of sefnicrystalline polymers by polarized light methods constitutes a large proportion of all light microscope studies on polymers.

Amorphous polymers, however, do not have any large-scale regular structures, so polarized light methods are confined, with this type of polymer, to examining orientation and/6r frozen-in stress. As outlined in Chapter 3, the degree and direction of orientation can be assessed from a section, both for crystalline and for amorphous materials. With transparent amorphous materials, however, birefringence can be assessed at least in a semiquantitative manner in whole mouldings. The interpretation of what is seen when whole mouldings are examined between crossed polars is full of pitfalls for the unwary since the colours observed are not solely dependent on the degree of orientation at a

111

112 B. P Saville

point. However, there is a lot of interest in the interpretation because of its obvious application as a simple non-destructive quality control test.

4.2 SPHERULITES

Spherulites are identified as such from their characteristic morphology. In polymers this can readily be observed when a thin section is examined between crossed polars. In general the spherulitic structure of polymers is not visible to the naked eye, so it has to be studied by polarized light microscopy. In their simplest form spherulites appear as circular birefringent areas showing a dark extinction pattern in the shape of a Maltese cross of which one arm is aligned with the vibration direction of the polarizer and the other arm is aligned with the vibration direction of the analyser (Fig.4.l). On rotation of the specimen this

FIG. 4.1. Section of acetal; crossed polars. Bar = 200,urn (X 150).

Polarized Light: Qualitative Microscopy 113

patterns remains stationary, the arms of the Maltese cross remaining parallel to the vibration directions of the polars. This shows that the section of the spherulite has circular symmetry which could be either radial or tangential. This view can be shown, by examination of a threedimensional specimen from more than one direction, to be a circular section of a spherically symmetrical entity.

These observations can be accounted for by a model in which identical birefringent units are arranged so that they radiate symmetrically in all directions from the centre of the spherulite. In the areas where the units are parallel to the vibration direction of either of the polars there will be extinction which will take the form of a cross. Indeed the fine structure of spherulites when examined under the microscope often shows a radiating branching fibrillar appearance as in Fig. 4.2. However, the relationship between these hypothetical units and the real structural entities of polymers such as chains, crystallites or lamellae is rather more complicated.

The spherulitic form of crystallization is not confined to polymers; it is found in such widely different types of materials as naturally occurring minerals and simple organic compounds. This has led to the hypothesis that the particular form of crystallization is dictated purely by the crystallization conditions which must have common features in all the cases where it is the preferred crystallization form.

Keith and Padden' have suggested that the distinguishing features of this type of crystallization, which are present in polymers and which they share with the other materials that crystallize in this form, are a high viscosity in the melt and a much slower rate of crystallization than for most other materials which prefer to crystallize as individual crystals.

4.2.1 Fine Structure Spherulites are not themselves crystals; they are aggregates of much smaller crystallites. The final diameter of the fully formed spherulite does not reflect the overall crystallinity of the polymer or the size of the individual crystallites but rather the conditions under which the polymer was crystallized. They are, however, a product of primary crystallization, not a rearrangement of already crystallized material, i.e. the crystallites are formed from the melt in the final positions they will occupy in the spherulite as it grows. That this is the case was demonstrated using poly(ethylene terephthalate)2 which can be quenched from the melt to an amorphous state by bringing its temperature quickly

114 B. P. Saville


below its glass transition temperature. It is possible, by quenching that material before crystallization is complete, to produce isolated spherulites in a non-birefringent matrix. Small areas of the spherulitic and non-spherulitic regions were examined using X-ray diffraction, by which means it was possible to demonstrate that only the spherulitic areas were crystalline. This proves that the spherulites were the crystalline portion of the sample, growing at the expense of the noncrystalline melt.

A distinguishing feature of a spherulite, which can be measured using a polarizing microscope, is its birefringence. This can be defined as

(4.1)

where nr is the refractive index in the radial direction and n t is the refractive index in the tangential direction. If the spherulite has its larger refractive index in the radial direction then it has a positive birefringence, whereas if the tangential direction has the higher index the spherulite has a negative birefringence. The sign of the spherulite birefringence is a characteristic of a polymer or, to be more accurate, the particular crystalline form of the material. The sign of the birefringence of a spherulite can most easily be determined using a sensitive tint plate which will add to the path difference of the spherulite or subtract from it, depending on whether the birefringent units in the spherulite are oriented parallel to the plate or perpendicular to it. This means that one pair of quadrants of a spherulite will be raised up the scale of colours and the other will be lowered. Therefore, with spherulites showing only first-order white, which is normal in a thin section, one pair will be raised to blue and the otherlowered to orange. With spherulites of the opposite sign the colours appearing in the quadrants are reversed.

4.2.2 Orientation of the Chains An important consideration, once the characteristic form of polymer crystallization was discovered, was how the basic unit, i.e. the polymer chain, is incorporated into the spherical structure. In certain cases the orientation of the polymer chains with respect to the radius of the spherulite could be deduced from the sign of the birefringence of the spherulite. If the sign of the birefringence in the spherulite is positive, this means that the larger refractive index lies along the radial direction, and if the sign is negative then the larger index lies at right angles to this

116 B. P. Saville

along the tangential direction. For most polymers the direction of highest refractive index coincides with the chain direction because most of the covalent bonds are aligned with this direction. Using this line of reasoning, it was deduced that the polymer chain direction in polyethylene spherulites was tangential, from the fact that the spherulites had a negative birefringence.

However, the relation between the molecular orientation and the sign of the birefringence is not so clear-cut for polymers in which the refractive index is high in one of the directions perpendicular to the chains, as is the case for polyamides and poly( ethylene terephthalate ).It is not necessary for this high index to be greater than that along the chain for the spherulite to show a positive birefringence. If it is higher than the average of the index along the chain and the lowest index of the molecule, the chains can still be oriented in the tangential direction but yet show a positive birefringence. This is because, in a given section of a spherulite with tangentially oriented chains, some of the chains will lie in the plane of the section and some will be perpendicular to it, so the tangential index will be an average of the index for the chain together with either one or both of the other indices. The radial index will be either one of the other two indices or an average of them, both depending on the degree of three-dimensional structure present. Several polymers do form either positive or negative spherulites, depending on the crystallization conditions. This could be due to either a slightly different orientation of the chain or a more regular three-dimensional ordering of the crystallite in directions perpendicular to the chain direction.

Fortunately more definite information on chain direction in spherulites can be obtained by recording X-ray diffraction patterns from small regions of spherulites using a very fine X-ray beam. In all cases studied it has been found that the polymer molecules lie more or less in the tangential direction. It was also found2 that in positive polyamide spherulites the hydrogen bonded planes were parallel to the radius but in the negative spherulites they were perpendicular.

The tangential arrangement of the polymer molecules appears to contradict the evidence of the fibrillar appearance seen in many spherulites. However, with the acceptance of the evidence that the basic crystallization form of polymers is the chain-folded lamellae, it can be seen that it is the lamellae that are radiating from the centre of the spherulite. Indeed, a mechanism of growth in which the successive chains are laid alongside one another is more to be expected since it is the mechanism of crystallization in the short-chain paraffins.


4.3 THEORY OF SPHERULITIC CRYSTALLIZATION

The crystallization mechanism that results in spherulites is different from that in operation in most other crystalline forms. In these each nucleus gives rise to a single crystal in which the material is laid down in successive layers with all the molecules having the same threedimensional orientation. A spherulite, although it also grows from a single nucleus, does so in the form of a multitude of crystalline fibrils oriented in all directions so as to fill a sphere centred on the nucleus. As the fibrils grow away from the nucleus they repeatedly branch so as to fill up the space available. The branching is said to be non-crystallographic because the angle between the branch and the parent fibre is not related in any way to the geometry of the crystal; the orientation of one particular axis of the individual crystallites remains more or less parallel to the radial direction of the spherulite.

The growth rate of spherulites is such that, under isothermal conditions, the radius increases at a constant rate. This means that, when spherulites that were nucleated at the same time meet during growth, the boundary between them is a straight line equidistant from the two adjacent nuclei; if the spherulites are nucleated at different times, the boundary is curved and further away from the nucleus that was formed first.

It is known from experimental evidence3 that the first product of spherulitic crystallization is a branched fibrillar structure and that the material between the fibrils crystallizes afterwards. This process which is known as secondary crystallization may carry on for long periods of time after the volume of the original melt has been completely filled with spherulites, and in some cases the spaces between the fibrils may remain amorphous. As the crystallizing temperature is increased the texture of the spherulites generally becomes coarser as the width of the individual fibrils increases.

The mechanism put forward by Keith and Padden l to account for spherulitic type growth in both polymers and non-polymers postulates a central role for impurities in the melt. With some organic compounds, such as resorcinol or malonamide, it is possible to convert the form of crystallization to spherulitic by the addition of a small proportion of an impurity. This gives a disproportionately large increase in melt viscosity together with a reduction in the rate of nucleation. It is found that the radial growth rate of the spherulites decreases with increasing impurity concentration.

118 B. P Saville

Polymer melts can be considered to be impure in that they contain a wide range of molecular weights together with molecules that are not stereoregular or those that contain chain branching. These molecules may crystallize less readily than the bulk of the material, orthey may not be incorporated into the crystal lattice at all. It has been shown experimentally that such molecules are rejected by the growing spherulites. For instance, an isotactic polypropylene deliberately diluted with atactic polypropylene,4 crystallization leads to spherulites with a coarse texture where the atactic material is concentrated between the fibrils. In polyethylene it is found that the low molecular weight fraction is segregated by crystallization at high temperatures.5 This fraction can be preferentially dissolved by hot solvent, to leave both intraspherulitic voids and voids at the outer edges of the spherulites.

It is considered that the rate of advance of the crystallization front is dependent not only on the diffusion of impurities, as defined above, away from the growth front. It is postulated that the growing crystal rejects the impurities, so making their concentration higher at the liquid/crystal interface than it is elsewhere in the melt. It is this impurity-rich layer, and in particular its depth, that is thought to give rise to the fibrous form of the spherulites and also to the noncrystallographic branching of the fibres.

The build-up of impurities next to the growing front inhibits crystallization in this region because of the greater degree of supercooling that would be needed to crystallize it. Depending on the thickness of the layer, any random surface instability can reach through this layer to the normal melt, beyond where crystallization could continue on this surface. Thus the material is preferentially crystallizing in a fibrillar form; in the process, impurity-rich melt is becoming trapped between the fibrils. Further lateral growth of the fibrils is then retarded by this concentration of impurity.

The degree of branching in a spherulite is thought to be controlled by the thickness of the impurity layer. The thinner it is, the easier it is for any surface imperfection to initiate growth on its own, giving rise to more frequent branching. This is thought to be responsible for the coarser textures found in spherulites crystallized at high temperatures (i.e. low supercoolings) compared with the finer textures found in those crystallized at large supercoolings. The effectiveness of the impurity layer at inhibiting crystallization is dependent on the degree of supercooling, which provides the driving force for crystallization, i.e. the greater the supercooling the less the effect of the impurity


concentration. Increasing the concentration of atactic polypropylene in isotactic polypropylene also leads to a coarser texture at a given temperature of crystallization.4

The impurity concentration in a polymer can be effectively increased by the production of low molecular weight species which occurs in degradation. Holding a polymer at high temperature for long times has been found to give rise to coarser textures.

4.3.1 Nucleation and Growth The formation of spherulites from the melt is governed by two separate mechanisms: nucleation and subsequent growth. These, however, can take place at the same time, further spherulites being nucleated whilst the original ones are growing, so the final size of spherulites formed depends on the relative rates of the two processes for the particular crystallization conditions existing at the time. Nucleation takes two main forms: homogeneous, in which the nucleus is formed out of the molten polymer, and heterogeneous, in which crystallization takes place on a foreign, pre-existing surface. The crystallization of polymers takes place at temperatures appreciably below the melting temperature.

The reason for the supercooling and the need for nucleation is believed to be that, although the formation of the crystalline phase is energetically favourable below the melting point, the creation of new surface in the crystal requires an extra amount of energy above this. At a given degree of supercooling of a polymer melt there is a critical size of nucleus, above which the crystal will continue to grow rather than be absorbed back into the melt. In the absence of any preformed nuclei, crystallization of the melt depends on random fluctuations within the melt providing nuclei above the critical size; the larger the critical size of nucleus required, the longer this process will take. The greater the degree of supercooling in the melt, the smaller will be the critical nucleus size; therefore the nucleation rate increases rapidly with decreasing temperature below the melting temperature. At much larger degrees of supercooling the nucleation rate decreases again owing to the increasing loss of mobility suffered by the polymer chains as the glass transition temperature is approached.

Heterogeneous nucleation takes place around existing particles of foreign material which are then incorporated into the crystal. This foreign material can be present as very low levels of contamination, or can be deliberately added to the melt to seed the crystallization. This type of crystallization can also take place on surfaces that are in contact

120 B. P. Saville

with the melt, leading to difficulties in the study of crystallization if these effects are not taken into account. The foreign surface acts by lowering the energy barrier to crystallization by reducing the amount of new free surface that has to be formed. One consequence of this is that if such a material is present in a polymer melt it will cause crystallization to take place in it at smaller supercooling than would be the case for homogeneous crystallization.

Only certain materials will act as nucleating agents for a given polymer, but it has not been possible to specify with any accuracy what type of material will so act. It has been found for polypropylene6 that a nucleating agent should possess a polar and an organic group. The number of active nuclei has been found to increase with the degree of

. . ~

"r_ .... ., "'-.. .,

•

FIG. 4.3. Section of linear low density polyethylene rotational moulding showing pigment; bright field . Bar = 50)lm (X800).


supercooling and also, at low concentrations, with the concentration of the foreign seed used.

Certain pigments can also act as nucleating agents, in particular copper phthalocyanine; this can lead to marked differences in mechanical properties between different colours of apparently similar products. Figures 4.3 and 4.4 show this effect in a rotationally moulded product where the polymer has been tumble-blended with dry pigment. Because of the lack of shear in this process, the pigment remains on the outside of the polymer granules. Figure 4.3 shows the pigment distribution in bright field and Fig. 4.4, from the same area between crossed polars, shows the resultant nucleation from this pigment.

A further phenomenon of nucleation is self-nucleation in which, as distinct from homogeneous nucleation, the polymer is nucleated by its

FIG. 4.4. Same area as Fig. 4.3; crossed polars. Bar = 50 J1m (X800).

122 B. P Suville

own crystals which have been grown previously. At temperatures just above the melting point of the polymer, some crystals can remain in the melt or can be trapped in cracks on foreign surfaces. These crystals have an effectively higher melting point than that of the bulk of the material; on cooling the melt, the crystals then act as nuclei. It is found experimentally in such systems that the supercooling necessary for crystal growth to occur in a given time or the time to crystallize at a given temperature is, over a limited temperature range, dependent on the degree of heating above the melting temperature of the bulk material. This shows that the number of nuclei decreases with increasing temperature above the melting point until a steady state is reached which depends on the number of heterogeneous nuclei present.

An important variant of self-nucleation is that which can be produced by orientation of the polymer chains in melts tha t are sheared. An extended polymer chain has effectively a higher melting point than that of the bulk material so that, at a given temperature below the melting temperature, an extended chain will have a larger supercooling, and thus there is a greater driving force towards crystallization. Moreover, the extended chain is closer to the final crystallized form of the polymer than is a random coil, so an enhanced rate of crystallization might be expected because of this. Figure 4.5 shows the type of structure known as row nucleation produced by this effect where a string of spherulites have their origin in the same region of sheared material. Sheared melts can also give rise to the shish-kebab type of structure in which the core is considered to be an extended chain fibre covered with overgrowths of chain-folded lamellae.

4.3.2 Crystallization The second factor that affects the final spherulite size is the rate of crystallization of the polymer which, like nucleation, is dependent on the degree of supercooling.

If the development of crystallinity in a polymer as it is cooled from the melt is followed by measuring, for example, the specific volume change (Fig. 4.6), it is found that the increase in crystallinity with time follows a characteristic pattern. An initial interval of time is observed in which there is no detectable increase in crystallinity, but once crystallization has started it proceeds at an accelerating rate until finally it appears to approach an equilibrium crystallinity value. However, the crystallinity can continue to develop further over long periods of time. The timescale over which these changes take place is strongly dependent on the

124

speCific volume

B. P Saville

Increasing

crystallization temperature

time

FIG. 4.6. Increase of crystallinity (decrease in specific volume) with time at different temperatures of crystallization.

temperature of crystallization relative to the melting temperature of the polymer, i.e. on the degree of supercooling. At temperatures just below the melting temperature the crystallization rate is very slow; as the temperature is lowered, the rate increases rapidly and eventually passes through a maximum.

At temperatures below this maximum the rate of crystallization becomes slow once again until the glass transition temperature is reached, at which point crystallization ceases. A polymer cooled quickly enough to this region would become glassy. However, in many polymers the crystallization rate is so rapid at temperatures below the melting temperature that it has been impossible experimentally to observe the maximum or any points below it in the curve of crystallization rate against temperature. Poly(ethylene terephthalate) is a material that can be quenched to a glassy state, and the full curve of crystallization rate against degree of supercooling has been observed.? This shows a broad maximum between 1600 e and 220 0 e, with the actual peak at 190°C. The experiments also showed that the density increases with temperature of crystallization, slowly until 220° e and at a greater rate of increase after this. If a material like poly( ethylene terephthalate) is quenched to a glassy state, crystallization will subsequently take place if the material is heated above its glass transition temperature; the rate at which this takes place is governed by the excess temperature above the transition temperature.


4.4 DIFFERENT TYPES OF SPHERULITE

So far we have considered only the model spherulite with the simple Maltese cross type of extinction pattern. However, in practice the appearance of spherulites varies from polymer to polymer and will often in a single polymer vary with the crystallization conditions. It is often the case with smaller spherulites that the individual ones are no longer identifiable as such, yet there is a 'spherulitic' texture apparent when the material is viewed between crossed polars. It is therefore possible to gain information about a polymer and the conditions under which it was cooled from the melt, by examining the appearance of the spherulites, provided that their variability has been carefully studied.

One of the more easily described features found in some spherulites is the appearance of bands or rings in the extinction pattern. These may be conveniently grouped into three types which can, however, merge into one another: (a) a pattern that, in addition to the Maltese cross, has regularly spaced concentric rings; (b) a pattern similar to that of (a) but with the concentric rings grouped in pairs; (c) a type in which the arms of the simple Maltese cross are replaced with zig-zag patterns of extinction. This is often associated with a concentric ring pattern, in which case the extinction cross zig-zags between the rings (Fig. 4.7). All variations between plain concentric rings and a zig-zag extinction cross can usually be observed, depending on the crystallization conditions.

The appearance of rings in the spherulite pattern is very much dependent on the rate of cooling from the melt; the same polymer can show either a simple extinction by itself or a ring pattern in addition as the conditions of crystallization are changed.

For example, rapid crystallization of high density polyethylene gives rise to spherulites showing a ring pattern, whereas slow crystallization of the same material gives spherulites with no rings.8 In the case of poly(ethylene terephthalate),9 crystallization temperatures of up to 1800 C give rise to spherulites with the usual extinction cross, above this appears the zig-zag form of cross, the arms of which elongate with increasing crystallization temperature until at 2390 C the ends of the adjacent zig-zags meet giving a pattern similar to that shown in Fig. 4.8. In the case oflinear low density polyethylene, quickly cooled material has a normal extinction cross (Fig. 4.9) whereas slowly cooled material shows concentric rings (Fig. 4.10). This variation in behaviour between the different types of polyethylene may be due to the difference in temperatures of crystallization experienced between bulk material that

126 B. P. Saville

S :::t g -

FIG

. 4.8

. T

hin

fil

m o

f po

ly(e

thyl

ene

tere

phth

alat

e);

cros

sed

pola

rs.

Bar

=

lO J

im (

X22

00).

cl' is"" i:l. It

t"-<

0'(;. ;::.. ~

~

." ~ ~. ~ <":

> ;:s 1'l {;

~ ...... !::l

128 B. P. Saville

FIG. 4.9. Section oflinear low density polyethylene, fast-cooled; crossed polars. Bar = 50.urn (X800).

has been allowed to cool slowly without quenching and material that has undergone a very slow isothermal crystallization at temperatures just below the melting point. Both would in certain circumstances be described as slow-cooling but one could also be described as fastcooling with respect to the other. It is also observed that the ring spacing varies with the temperature of crystallization in polyethylene,IO the spacing being larger at lower degrees of supercooling and becoming increasingly smaller as the supercooling is increased.

There are a limited number of possible explanations for the banded type of extinction pattern:

(1) The material that gives rise to the dark rings may have a principal


FrG.4.10. Section of linear low density polyethylene, slow-cooled; crossed polars. Bar = 50.urn (X800).

axis parallel to one of the polars, i.e. they are at extinction for the same reason as the material that gives the Maltese cross. This explanation can be examined by viewing the spherulites in circularly polarized light (Fig. 4.11) which removes any directional effect of the polars (see Chapter 3). It can be seen that the extinction cross visible in Fig. 4.12 is removed but the concentric rings remain.

(2) The material in the dark rings has its optic axis, or axes, parallel to the direction of the incident light. For a uniaxial polymer this would generally mean that the polymer chains were oriented perpendicular to the section at that point. For a biaxial polymer there would be two orientations of the chains that could produce extinction when either of the two optic axes was parallel to the incident light. This is usually

FIG

. 4.1

1.

Th

in f

ilm

of

high

den

sity

pol

yeth

ylen

e; c

ircu

larl

y po

lari

zed

ligh

t. B

ar =

20.

um (

X 1

3(0)

.

-w o P:l ~

~ '" ~


8' M -X '-'

132 B. P. Saville

considered to be the explanation for the double extinction rings. (3) The material in the dark rings could be disordered, i.e. non

birefringent or isotropic.

Explanation (2) is the generally favoured one,lI although there is some support for (3).8.12 As the polymer chains in a spherulite are arranged more or less tangentially in the form of lamellae, this would imply that the lamellae had a regular twist as they grew in the radial direction, giving rise to two extinctions for every 360° of rotation in the uniaxial case or four for biaxial crystalline material.

4.5 SPHERULITIC FORMS OF POLYPROPYLENE

A number of polymers form different types of spherulites, such as with positive or negative birefringence, depending on the crystallization conditions. Polypropylene is a particularly prominent example, for which four different types of spherulite have been identified13 in samples isothermally crystallized at different temperatures below the melting point.

Type l. This is the predominant form at crystallization temperatures below about 134°C. It has a positive birefringence of magnitude 0·003. The extinction pattern is a simple Maltese cross.

Type II. This grows preferentially at crystallization temperatures above 130° C. It is distinguishable from type I by its birefringence which is negative and of magnitude 0·002.

Type III. These spherulites appear at crystallization temperatures below 128° C and their formation seems to be favoured by rapid cooling from the melt temperature. Even under the most favourable conditions they usually only comprise a small fraction of the total volume and appear scattered throughout the material as shown in Fig. 4.13. They are readily distinguishable by their much greater negative birefringence which has a magnitude of 0·007.

Type IV. This is similarly isolated in its appearance but is formed between 128° C and 132° C. It is a ringed spherulite but with a negative birefringence higher than that of type I.

Microbeam X-ray studies of examples of all four types of spherulite l4

showed that the crystalline forms of both types I and II were the same but that types III and IV though similar to one another had a different crystalline structure.


8 ::t

8 ......

~ ~

'0 0..

"0 o '" '" e u Q) ~ o

~ e ~ '0 0.. ..... o ~ o

'B o

__ , ~,---,..=- lJ:J

<'"i

~

d ~

134 B. P. Saville

8 :::t

§ ....

136 B. P Saville

This was confirmed later on samples where the different crystalline forms could be obtained substantially free from one another. The second crystalline form occurring in types III and IV spherulites was termed the f3 form to distinguish it from the a form occurring in types I and II. It was also confirmed that the f3 form is converted into the a form at temperatures above 130° C; the higher the temperature the faster is this conversion. This explains why the growth of the f3 form is favoured by rapid cooling to below about 130° C. Experiments with quenched material showed that the f3 form was most prominent at quench temperatures between 100° C and 120° C; below 90° C and above 130° C mainly the a form is produced.

In practice only two types of spherulite are widely observed, type I and type III. Type III spherulites are found dotted amongst the predominant type I, their number and position in a moulding being a strong indicator ofthe crystallization and shear history of the moulding. Figure 4.l4 shows a fast-cooled polypropylene moulding; compare this with Fig. 4.l5 which shows slow-cooled polypropylene, and note the lack of the bright f3 form spherulites in the outer skin of the fast-cooled sample. It is interesting that in both cases most of the bright spherulites are concentrated in a band inside the skin; this is presumably the region that is rapidly cooled to below 130° C but not below 90° C and where shear forces during injection are highest. The interior of the moulding in both cases is more slowly cooled.

4.6 EFFECT OF PROCESSING ON SPHERULITES

Since both spherulite nucleation and growth are affected in the same way by supercooling, it is not possible to predict the effect of different cooling rates on spherulite size. However, in practice it is recognized that fast cooling gives rise to small sphemlites and slow cooling gives large ones. This can readily be seen at the surface of a fast-cooled moulding (Fig.4.l6) where the size decreases towards the skin compared with a similar slow-cooled moulding (Fig. 4.17) where the size does not change. Because of the poor heat conduction of polymers, the interior of all but thin-walled mouldings are generally slow-cooled.

Polymers crystallized with large spherulites have a higher crystallinity but are more brittleY This is thought to be due to a greater concentration of defects, including voids, at the edges of the large spherulites, which

138 B. P. Saville

x '-'

S ::t o o N

II .... ~ ~

"0 (1)

'0 o u ~ o til


have had more time to order themselves. It is therefore generally recognized that, as a smaller spherulitic structure is to be aimed for in manufactured products, this can be achieved by the use of heterogeneous nucleants in some polymers. Quenching or low melt temperatures, as a means of controlling spherulite size, can easily be carried too far, resulting in high molecular orientation. In Fig. 4.18 the outer skin is highly birefringent where the polymer has been quenched while still flowing, thus freezing-in the orientation. This resulted, in this particular component, in a brittle skin easily cracked in the direction of orientation during assembly. Orientation caused by shearing of the melt against a quenched outer layer can give rise to row nucleated structures as in Fig. 4.5.

Use can be made of such information to help interpret more complicated situations of flow and thermal treatments.

Figure 4.19 shows a cross-section of half of a hot-plate weld in medium density polyethylene. On the right-hand side the large ringed spherulites are part of the original slow-cooled material. The band of sheared material in the centre is the line up to which the material was melted prior to the weld being made. As the weld is made, the molten polymer is squeezed out of both sides of the weld, and along that particular line the material is at such a temperature that it freezes immediately still in the sheared form. On the left-hand side the material is part of the actual weld, but it has had time to relax during cooling which has been somewhat faster than the original, which is why the spherulites are smaller. The sheared zone is the weakest part of the weld since it is here that the molecules are parallel to one another, so giving a plane of weakness. 16

A spherulitic phenomenon that is sometimes encountered, particularly with acetal, is the 'comet' spherulite as shown in Fig. 4.20. This is a product of the interplay between nucleation rate and growth rate as influenced by the temperature. Here, where the outside has been cooled more quickly than the interior, the rate of nucleation falls off sharply with increasing temperature as we go inwards from the outside. This leads to more growth occurring on the high temperature side of the nucleus. Figure 4.21 shows how a change in spherulitic structure points to a difference in processing conditions. The large spherulites on the outside of this rotationally moulded polyethylene component were found, by subsequent infrared spectroscopy, to be due to oxidized polymer which was reponsible for the brittle failure of this part.

140 B. P. Saville


A o 00 N X '--'

142 B. P. Saville

FIG

.4.2

1.

Sec

tion

of

oxid

ized

med

ium

den

sity

pol

yeth

ylen

e; c

ross

ed p

olar

s. B

ar =

50

pm

(X

540)

.

~

Ei [ r a;;'

;::-~

~

I:l r ~ ~ ~ .g '" .... e

144 B. P. Saville

4.7 SMALL ANGLE LIGHT SCATTERING

A frequent requirement in experimental studies relating polymer structure to properties is a measurement of average spherulitic size, particularly from selected areas of a specimen which can then be related to the crystallization conditions at that point. Unfortunately, in many fast-cooled specimens, at least part of the structure is too small to be easily measured using the light microscope. Small angle light scattering 17. 18 is a method of indirectly measuring spherulites in this size range, producing an average result for all the spherulites in a restricted area.

In order to obtain the light scattering pattern, a thin sample of the polymer is irradiated with polarized monochromatic light from a laser

FIG. 4.22. Small angle light scattering pattern from polyethylene.


source. The light is scattered by the spherulites in the sample and then passes through an analyser, after which the scattering pattern can be recorded on a photographic plate or by some other suitable means. The patterns are usually recorded with the analyser perpendicular to the direction of vibration of the incident light (Hv scattering) when a fourlobed pattern as shown in Fig. 4.22 is observed. Each of the four lobes has an intensity maximum near its centre, and it is the angular separation of diametrically opposite maxima that is inversely related to the spherulite size.

Using a medium power objective lens and with the field and aperture irises almost closed, it is possible to obtain such a low angle light scattering pattern in the back focal plane of the objective lens.

4.8 MOLECULAR ORIENTATION

In the study of non-crystalline polymers, polarized light can no longer reveal the microstructure, as is the case for crystalline polymers with their spherulites. However, it can be used to examine any orientation that may be present in products made from this type of polymer, as indeed it can for crystalline polymers. It is more usual for use to be made of the information revealed in a semi-quantitative manner. The direction of the orientation can be determined by rotation of the specimen between crossed polars and observing the extinction patterns. Its magnitude can be assessed by comparing the colour in white light with a Michel-Levy chart. This is sufficient to enable specimens to be compared with one another. The absence of any quantitative information on what is an acceptable level of orientation in a given polymer makes absolute measurements largely meaningless.

In the case of the examination of thin sections containing molecular orientation the interpretation of the observations is straightforward. The section can be considered thin enough so that none of the refractive indices changes through the thickness. The direction of cutting of the section can also be altered to coincide with the direction of the orientation when the maximum birefringence will be obtained. This is an important consideration for comparisons must be made using sections cut from the same relative positions; the observed birefringence will fall away as the angle between the direction of cutting and the direction of the orientation in the component increases.

In the case of whole transparent mouldings or extrudates which can

146 B. P. Saville

be examined using only two polaroid sheets, the problems of interpretation are much greater. One of the reasons for this is that the orientation changes through the thickness of a moulded part. It is generally greatest in, or just below, the faster cooled exterior skin and reduces or even reverses in direction towards the centre which has had time to relax as it cools. The optical path difference observed, when the whole of the wall thickness is examined between crossed polars, is the sum over the thickness of the moulding. Therefore it is possible to underestimate the orientation when it is present as a thin skin, but the mechanical effect of an oriented skin has a far more serious effect in making a part brittle than orientation elsewhere, as shown in Fig. 4.18.

Another difficulty arises where there is biaxial orientation such as is deliberately introduced when making oriented blow-moulded bottles. What is observed when such a sample is examined between crossed polars is the balance between the two orientations. This is similar to the case for biaxially oriented film mentioned in Chapter 3. Normal examination between crossed polars can only give information about the differences in the two refractive indices in the plane of observation. This is illustrated in Fig. 4.23 which shows the neck of a biaxially oriented bottle; there is an extinction band across the neck of the bottle where the balance of the orientation changes. In the neck of the bottle the overall orientation is parallel to the length of the bottle whereas below this in the body of the bottle the overall orientation is circumferential, caused by the blowing-up of the bottle predominating in this part.

It is possible to simplify some of the interpretational difficulties by using monochromatic light as in Fig.4.24 where the interference colours are replaced by black lines showing whole wavelength path differences, so giving a form of contour map of the path differences. This can be coupled with observation in circularly polarized light which removes any extinction bands parallel to the polars.

In certain cases an observed birefringence may be ascribed to the photoelastic effect. Most polymeric materials which are normally isotropic, because of random molecular orientation, can be made optically anisotropic by the application of mechanical stress. In going from tension to compression the material will change from positive uniaxial to negative uniaxial or vice versa depending on polarizability considerations. In both cases the effective optic axis is the stress direction. The birefringence at any point is proportional to the principal stress difference (0', - 0'2)' In the elastic region of the material the

Polarized Light: Qualitative Microscopy

birefringence can be related to the stress by

n1 - n2 = C(0"1 - 0"2)

147

(4.2)

where C is the stress-optical coefficient which has usually to be found experimentally.

Distinction between stress (or strain) optical effects and those due to molecular orientation can present practical problems and may be overlooked. One possible solution is to raise the temperature of the specimen under observation to just above its glass transition temperature to allow stress relaxation. However, it should be remembered that the

FIG. 4.23. Biaxially orientated PVC bottle neck; crossed polars.

148 B. P. Saville

birefringence, whether due to stress or molecular orientation, may be temperature dependent, so changes produced by elevated temperatures should be interpreted with care.

FrG.4.24. Polystyrene moulding; crossed polars, monochromatic light.


REFERENCES

1. Keith, H. D. & Padden, F. 1.,1 Appl. Phys., 34 (1963) 2409. 2. Keller, A, 1. Polymer Sci., 17 (1955) 351. 3. Hoskins, S., Meinecke, E., Powers, J., Stein, R. S. & Newman, S.,1. Polymer

Sci., A3 (1965) 3041. 4. Keith, H. D. & Padden, F. J.,J Appl. Phys., 35 (1964) 1270. 5. Winram, M. M., Grubb, D. T. & Keller, A, 1. Mater. Sci., 13 (1978) 791. 6. Binsbergen, F. L., Polymer, 11 (1970) 253. 7. Cobbs, W. H. & Burton, R. L.,1. Polymer Sci., 10 (1953) 275. 8. Low, A, Vesely, D., Allen, P. & Bevis, M.,J Mater. Sci., 13 (1978) 711. 9. Keller, A, 1. Polymer Sci., 17 (1955) 291.

10. Lindenmeyer, P. H. & Holland, V. F.,1. Appl. Phys., 35 (1964) 55. II. Geil, P. H., Polymer Single Crystals. Interscience, New York, 1963. 12. Keith, H. D. & Padden, F. 1.,1. Polymer Sci., 31 (1958) 415. 13. Padden, F. J. & Keith, H. D.,1. Appl. Phys., 30 (1959) 1479. 14. Keith, H. D., Padden, F. 1., Warter, N. M. & Wyckoff, H. W.,J. Appl. Phys., 30

(1959) 1485. 15. Way, J. L., Atkinson, 1. R. & Nutting, J., 1. Mater. Sci., 9 (1974) 293. 16. Oliveira, M. J. & Hemsley, D. A, Brit. Polymer 1., 17 (1985) 269. 17. Stein, R. S., Optical methods of characterizing high polymers. In Newer

Methods of Polymer Characterization, ed. B. Ke. Interscience, New York, p.155.

18. Haudin, J. M., in Optical Properties of Polymers, ed. G. H. Meeten. Elsevier Applied Science, 1986.

5

Modulation Contrast and Differential Interference Contrast Techniques

R. HOFFMAN

Modulation Optics, Greenvale, New York, USA

5.1 INTRODUCTION

If we compare the images of an isotropic phase object using the modulation contrast system (MCS) and the differential interference contrast (DIC) system in Fig.5.1(a) we note that the images seem equivalent in that they both appear three-dimensional. This is because each system produces an image where one side of the object appears bright and its opposite side is dark relative to a grey background. When images are shaded in this manner they appear three-dimensional. Now compare the images of an anisotropic phase object in Fig. 5.1(b) and note that the MCS produces an image that appears more threedimensional than the one produced by DIe. This chapter will provide information about the equipment needed to produce such images, explain why the images appear as they do and describe how to apply this information to polymeric specimens.

5.2 GENERAL PRINCIPLES

What are the physical characteristics of objects that are detected and displayed by these microscope systems? The MC and DIC systems detect the optical gradients and convert them to intensity variations. Since the eye can only detect color and intensity variations, detail in non-absorbing (phase) objects, such as in a piece of transparent polymer, is normally invisible. The optical gradient occurs at the curved

151

152 R. Hoffman

(a)

(b)

FIG. 5.1. (a) Comparison photomicrographs showing the modulation contrast system (left) and the differential interference contrast system (right) views of an isotropic object. Note the three-dimensional appearance of each image. (X400.) (b) Comparison views of an anisotropic object showing that the modulation contrast view (left) appears three-dimensional while the differential interference

contrast view (right) does not. (X400.)

or sloped portion of an object and is the rate of change in the optical path difference (OPD) between the object and its surroundings. Figure 5.2 shows the representation of curved boundaries as optical gradients. The ray in Fig. 5.2 is proceeding in a direction indicated by the arrow. At the object boundary the ray is deflected through an angle cp, the optical

Modulation Contrast and Differential Interference Contrast

I

4~B:"""~:" :L ~_~e _____________ Incident

wove front

r Incident direction

(a)

Point of tangent

(b)

153

FIG. 5.2. (a) Diagram of a cUlVed transparent phase object showing continuously changing gradients represented as 'prisms'. (b) Diagram showing the deflection of a ray at the boundary gradient. (From Ref. I, reproduced by permission of the

Journal of Microscopy.)

gradient, and proceeds in the direction indicated by the arrow after the boundary. The tangent to the boundary establishes a, the geometric boundary gradient. If we idealize an object as in Fig. 5.3, opposite gradients appear on opposite sides of the object. The direction of deflection of the ray is dependent on the relative refractive index and analyzed according to Snell's law. I An opaque object observed using a reflected light microscope can also be considered a phase object, where cp = 2a.

The optical gradient is the primary feature detected by MC and DIC systems. The magnitude and direction of the optical gradient, cp, is dependent not only on its boundary angle, a, but also on the difference in refractive index (No - N m), where No andN m are the refractive index of the obiectand the media. respectively. For small anglescp = a(No - Nm).

154 R. Hoffman

3 4 2 N,>N2 N,<N2 0 N,<N2 N,>N2

A' B'

N2 N,

A B

FIG. 5.3. Diagram of a phase object showing the pairing angles of deflection (I/J) relative to the boundary angle (a) and the relative refractive indices N1 and N2 •

Note the opposite deflections of the pair of rays I and 2. (From Ref. I, reproduced by permission of the Journal of Microscopy.)

An object with more than one refractive index, i.e. an anisotropic material, will have more than one optical gradient. The direction of deflection of the light wave for each optical gradient is the same if the two refractive indices are either greater than or less than that of the surround. If the refractive index of the surround is between the maximum and minimum refractive indices of the object, the gradient magnitude and direction will be the resultant of the two refractive indices. Thus, if the signs of both gradients are the same the contrast will be greater than if one is opposite to the other.

Later we will show how the use of one polarizer affects the image of polymer specimens, which are generally anisotropic materials. Other important physical features affecting the image of the object are its size, edges and regularity of spacing. The effect of these features on the final image depends on the wave nature of light, the diffraction process and the relative coherence of the light just prior to its interaction with the object. Since the mid-wavelength in the visible spectrum is about 0·5.um, physical features around this size create strong diffraction effects. The image of these features depends on the recombination (interference) of the diffracted waves at the image plane. The visibility of this interference process is dependent on the relative coherence oflight at the moment of interaction with the object. 2 The greater the coherence the greater the visibility of the interference process, and thus the more visible are the images of the diffraction sites in the object. In a microscope, the coherence of the light that reaches the object plane is controlled by the aperture of the condenser system.3 These ideas

Modulation Contrast and Differential Interference Contrast 155

correspond to the common observation that as the condenser aperture is decreased the visibility of edges and particles of phase objects is increased, although resolution is reduced.

Special contrast systems are needed to view the optical gradient in phase objects, both in transmitted and in reflected light.

5.3 THE MODULATION CONTRAST SYSTEM

5.3.1 Optical Details of the System A bright field microscope can be modified to a modulation contrast system by adding a special slit aperture below the condenser and a rotatable polarizer (P I) below the slit aperture (Fig. 5.4). A 'modulator' is placed in the objective at a plane conjugate with the slit aperture. A different size slit aperture is needed for each objective magnification.

Eyepiece <===> G B

Modulator . , -([) Objedive <===>

o

Specimen r=== ]

Condenser

Sli

P, r/ZZI?I??????IZZ?...,

FIG. 5.4. Schematic diagram showing the components of the modulation contrast system added to a bright field microscope. The left plan view shows the modulator regions: dark (D), grey (G), bright (8). The right plan views show the slit and ~it image correctly registered and superimposed on the modulator. PI and P2 are polarizers. (From Ref. 1, reproduced by permission of the Joumal of

Microscopy.)

156 R. Hoffman

The modulator has three regions: a small dark region (D) at one edge with a light transmission of less than 1%; a narrow grey region (G) of approximately 15% transmission; the remainder of the conjugate plane which is a bright region (B) of 100% transmission. There are essentially no phase changes introduced by the modulator. The slit aperture is a slit partially covered with a polarizer (P2 ) and is imaged at the modulator plane. When PI and P2 are crossed the effective slit is narrow and its image registered in the grey region of the modulator. The portion of the slit image controlled by the polarizer is registered in the bright region of the modulator. Rotating PI varies the effective width of the slit and thereby varies the spatial coherence and contrast. The slit aperture and the grey region of the modulator are offset from the optic axis to permit maximum resolution. When the slit is removed, the modulation contrast objective can be used for bright field, dark field, polarization and fluorescence modes. The transmission MCS can be used in combination with reflected fluorescence techniques.4 Other configurations with the slit aperture will be described in the section on polarized light with modulation contrast.

The modulation contrast microscope is a system based on five principles which are demonstrated by considering an optical gradient to be a small prism.

(1) A prism deflects a ray of light. Figure 5.5(a) represents a typical microscope condenser-objective combination. A slit is located at the front focal plane of the condenser and its image is formed at the conjugate plane, the back focal plane of the objective. Figure 5.5(b) illustrates the deflection oflight ray 2 when a prism is placed in its path.

(2) The slit image, 2 in Fig. 5.5(b) is shifted by light ray 2 passing through a prism while light ray 1, passing around the prism, forms a slit image that is not shifted. This is indicated by the two slit images in Fig. 5.5(b).

(3) Shifted slit images pass light through different portions of the modulator. When a specially designed filter, the modulator, is placed at the back focal plane (Fig. 5.5(c», the light of the shifted slit image 2 passes unattenuated through the bright region while the light from the non-shifted slit image 1 is attenuated to 15% of its original intensity as it passes through the grey region.

(4) The intensity of the image of the optical gradient is controlled by the transmission factor of that portion of the modulator through which the rays pass. After passing the bright region of the modulator, the light forms a bright image of the optical gradient at the real image plane, and


Image plane

Slit image = Bock focol plane

.c:: :> Objective

I f Specimen

-=- ~ Condenser

- _ Front focal plane (slit)

(a)

Bright imoge of B optical gradient r===J G

o tG fB I 2 Shilted b I Modulator

EJ = =--'slit image

f f .c:: :0-

I Deflected roy I ..:::::::::J ..:::::::::J

It 2t t t ~

- - --(b) (c)

FIG. 5.5. Schematic diagram of a condenser-objective combination. (a) Paraxial rays form a slit image on the optic axis. (b) A deflected ray 2 leaving an optical gradient (prism) forms a slit image shifted off the optic axis. (c) The slit images are shown passing through different portions of the modulator to form a bright image of the optical gradient against a grey background. The lengths of the arrows indicate relative intensity. 0, dark; G, grey; B, bright. (From Ref. 1,

reproduced by permission of the Journal of Microscopy.)

the light that did not pass through an optical gradient passes through the grey region to form the grey background in the image plane (Fig.5.5(c». Therefore the prism gradient image is brighter than the background and thereby becomes more visible. An opposite gradient causes light to be highly attenuated by the dark region of the modulator. This creates a dark gradient image. If no modulator were present (Fig. 5.5(b », the intensity of the shifted and non-shifted slit images would be equal when they arrived at the real image plane. The prism would be essentially invisible because the intensity of the gradient region of the specimen and background would be the same. Experience with oblique illumination would seem to contradict this statement. In fact, the edge of the back focal plane of the objective always acts as a filter to cut off the light that has been shifted by a large optical gradient, and the image of that gradient therefore appears dark.

(5) The diffracted waves pass through the bright region of the modulator to interfere at the primary image plane and form the image of fine detail and edges.

158 R. Hoffman

5.3.2 Non-ideal Specimens Rounded objects have continuously varying optical gradients. Each optical gradient creates a different angle ·of deflection, causing the image of the condenser slit to cover a range of positions in the back focal plane of the objective. Each slit image must pass through a different portion of the modulator. Many images of the slit appear simultaneously: one for each gradient and one for each point in the surround. The slit images for the surround are all superimposed and appear as a slit image registered in the grey region.

Figure 5.6 shows a series of optical gradients designated as prisms shifting the image of the slit relative to the modulator grey region. The intensity of the light before the modulator is the same for each optical gradient since the object does not absorb light. The length of the arrow after the modulator indicates the relative intensity for each gradient that will reach the image plane. Gradient 'c' is zero and represents the flat regions of objects as well as rays passing around the object through the surround. The slit image is registered within the grey region of the modulator which absorbs 85% of the light and passes 15% to the image plane to create a grey background and flat regions of the object. Gradient 'd' shifts the image of the slit so that one-quarter of its area passes the bright region of the modulator which transmits 100% while three-quarters of the slit image area still passes through the grey region

Pion view

Modulatar .;-:-;.:3 0 tG B t .ox.:.]

slit image .:··.·.·.1 ,"",-:.:1 -·:·;·.·,1 = = = = = \ \ r I / Prism ~ ~ C:::.::::... ~ =

t t l r (a) (b) (c) (d) (e)

FIG. 5.6. Diagram illustrating how the modulator produces intensity variations within a range of optical gradients. Length of arrow after modulator indicates intensity. The sum of the intensities of areas 1 and 2 is the intensity of the gradient after the modulator. (From Ref. 1, reproduced by permission of the



of the modulator. The intensity after the modulator is the sum of the intensity of each slit image area, i.e. areas 1 and 2 in the plan view. The intensity in the image plane for gradient 'd' is greater than that for gradient 'c'. The image intensity of an opposite gradient, 'b', is less than the background value. The maximum change of intensity occurs when the direction of the plane of the gradient is parallel to the length of the grey region.

The image of the slit is always shifted in the direction normal (perpendicular) to the direction of the plane of the gradient. Figure 5.7 illustrates the direction of the plane of the gradient and the direction of shift of the slit image. In Fig.5.7(a) the relative orientation permits maximum detection of the optical gradient. In Fig. 5.7(b) the specimen is rotated 90°, resulting in minimum detection of the gradient. Note in Fig. 5.7(b) the shifted slit image remains in the grey region.

5.3.3 Anisotropic Specimens Modulation contrast can be used to image each optical gradient separately in anisotropic materials such as polymer films and fibres. A

@ Modu la1or plane ";i'i~'

I

(a) (b)

~ / ~ / /~~<>;~"'~. ~~

op .. ,-- I I Op1 ic o.is

FIG. 5.7. Perspective diagram showing the direction of shift of the slit image at the modulator plane in relation to the direction of the plane of the optical gradient. The gradient in (b) is rotated 90° to (a). (From Ref. 1, reproduced by

permission of the Journal of Microscopy.)

160 R. Hoffman

slightly different version of the components shown in Fig. 5.4 is required. The polarizing portion of the special slit aperture needs to be removed or covered with an opaque material. If removed it needs to be replaced by an opaque material such that the original slit width (the width of the non-polarized portion of the slit) is retained. This part is imaged on to the grey region of the modulator. Rotating the polarizer that was placed before the slit isolates the direction of vibration in the anisotropic material. Figure 5.8 shows the effect that rotating the polarizer has on the image of an anisotropic grain. The photomicrograph shows an object where the refractive index of the medium is between that of the minimum and maximum indices of the object. In Fig. 5.8(a) the polarizer is parallel to one permitted direction of vibration in the crystal and shows the right side of the anisotropic crystal as bright. In Fig. 5.8(b), with the polarizer 90° to that used in Fig. 5.8(a), the right side is dark.

If an analyzer is added after the objective in the above configuration, the arrangement becomes a standard polarizing microscope combined with the MCS. With this combination the polarizing microscope detects the birefringence while the optical gradients of structures within or on the surface of the object are revealed by the MCS. Composite materials containing isotropic and anisotropic components may be observed with modulation contrast under parallel polars as shown in Fig. 5.9. With

(a) (b)

FIG. 5.8. MCS view of an anisotropic object using a polarizer before the object to select the orthogonal preferred directions of vibration. The refractive index of the medium is between the maximum and minimum refractive indices of the object. Note alternation of shading when the polarizer in (b) is rotated 90° to the

position in (a). (X330.)

Modulation Contrast and Differential Inteiference Contrast 161

crossed polars (Fig. 5.10) the optical gradients are revealed in only the anisotropic material when its orientation is not at extinction. The optical gradients in the isotropic material are not visible, nor are any other parts of the isotropic material except for the edges where there is some depolarization due to scattering. However, between parallel

FIG. 5.9. MCS view of cross-section of a composite of isotropic and anisotropic material under parallel polarizers. The color of the anisotropic material is blue.

Isotropic material is visible (see Fig. 5.10). (Xno.)

FIG. 5.10. MCS view of cross-section of a composite of isotropic and anisotropic material under crossed polarizers. The color of the anisotropic material is

yellow. Isotropic material is not visible. (Xno.)

162 R. Hoffman

polars the optical gradient structures in both the isotropic and anisotropic components are revealed. The optical gradient structures can only be detected when the zero-order energy passes through the optical system. Under crossed polars this energy is zero for isotropic materials and non-zero for anisotropic materials when the specimen is not at extinction.

5.4 DIFFERENTIAL INTERFERENCE CONTRAST

5.4.1 Isotropic Specimens Figure 5.1(a) shows the similarity of images of isotropic substances viewed with MCS and Ole. Note again that one side ofthe image is dark while its opposite side is bright against an intermediate value of the background. In the DIC microscope as in MCS, the optical gradients are also converted to intensity variations in the image plane; the sign of the gradient is preserved and the system is directional.

There are several optical configurations that can be used to create a DIC system. Generally these are based on a beam-splitting interferometer using birefringent elements between crossed polarizers.5 Currently the most common system, due to Normaski, uses two Wollaston prisms as shown in Fig. 5.11. One prism is effectively located at the front focal plane of the condenser system (WI) and the other after the objective in its back focal plane. Practical design considerations may mean that the prisms cannot be physically located at the appropriate positions. In these cases the prism design is modified so that their function is nevertheless optically correct. The Wollaston prism after the objective (W2) is called the compensator and is movable normal to the optic axis.

p

FIG. 5.11. A schematic diagram of the Nomarski differential interference contrast microscope showing the Wollaston prisms WI and W2 between crossed polarizers PI and P2 (C, condenser; OJ, objective; P, specimen; O2, ocular; Fo , back focal plane of objective). (From Ref. 1, reproduced by permission of th~


Modulation Contrast and Differential Inteiference Contrast 163

This motion controls the intensity and color in the image plane. Also the compensator splits or shears the imaging waves, in one direction, through a distance that is effectively within the resolution of the objective being used. When W2 is moved relative to WI the color of the field varies in accordance with Newton's interference color sequence and depicted in the chart for crossed polars shown in Fig. 5.12. The

First order

Second order

Path difference

(/-1m)

0'000 0'040 0'097 0'158 0'218 0·234 0'259 0·267 0'275 0'281 0·306 0'332 0·430 0'505 0'536 0'551

0·565 0'575 0'589 0·664 0·728 0'747 0·826 0·843 0'866 0·910 0·948 0'998 HOI 1-128 1·151

Interference Intensity colors between

crossed polarizers

Black Iron-grey Lavender-grey Greyish blue Clearer grey Greenish white Almost pure white Yellowish white Pale straw-yellow Straw-yellow Light yellow Bright yellow Brownish yellow Reddish orange Red Deep red

Purple Violet Indigo Sky-blue Greenish blue Green Lighter green Yellowish green Greenish yellow Pure yellow Orange Bright orange-red Dark violet-red Light bluish violet Indigo

FIG. 5.12. Newton's color scale for crossed polars, showing damped sin2

intensity curve for white light.

164 R. Hoffman

colors generated are essentially the same whether W2 is moved right or left from a central position. Associated with the color changes are intensity changes as shown by the curve in Fig. 5.l2. When W2 exactly compensates WI the field is dark and is called the zero-order position.

With a specimen in the field of view of the microscope system the DIC is designed to image the optical gradient. However, the DIC image does not always appear three-dimensional. When the compensator is set for a background of order zero, ! or 1, an object with small optical gradients does not assume the three-dimensional appearance (Fig. 5.13). Remember the perception of three-dimensions depends on one side of the image appearing bright while its opposite side appears dark against an intermediate grey value in the background. The setting of the compensator is thus critical in producing an image that appears threedimensional (Fig. 5.14).

In the phase gradient object shown in Fig. 5.l(a) the refractive index of the object is less than that of the surround; thus the wavefront emerging from the object (not considering diffraction from the edges) is that shown in Fig. 5.l5(b). We can arbitrarily assign the right optical gradient as positive and the left gradient as negative. The connection between the optical gradient and the OPD is through the expression

OPD = ¢d (5.1)

where ¢ is the optical slope or optical gradient and d is the effective shear distance produced by the compensator. The gradient is considered small.

Consequently, since opposite sides of phase gradient objects have opposite gradients, one side will produce a net OPD that is greater than the background OPD and the opposite side will produce a net OPD less than the background OPD. The background OPD is set by the compensator. Figure 5.13 shows a spheroidal object at different compensator settings. Figure 5.14 shows the intensity distribution across the image at various compensator settings. For example, when the background OPD is set at orded(OPD = 0·200 pm) and a small gradient object which has an OPD ofO·100 pm (eqn (5.l)) on each side is placed in the field of view, one side will have a net OPD of 0·100 pm while the opposite side will have a net OPD = 0·300 pm. Referring to the intensity curve in Fig. 5.l4(e) one side is darker than the background while the other side is brighter than the background. Thereby resulting in the perception of a three-dimensional image. If that same object were placed at order! approximately the same intensity would appear on each side of the

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optical gradient (cf» and arbitrarily designated as plus and minus.

object thereby resulting in no perception of three-dimensionality (Fig.5.l4(c)).

5.4.2 Anisotropic Specimens The intensity distribution in the polarization-based DIC image of an anisotropic object is not the same as for an isotropic specimen. The differences in the images can cause severe problems of image interpretation and occur regardless of the compensation setting. Figure 5.1 (b) shows such an image. Under DIC the image does not appear threedimensional, and note that the intensity appears similar on opposite sides. The sign of the gradient is not preserved and the image appears similar to that when viewed in a polarized light microscope. Sometimes weakly birefringent material, such as certain polymer spherulites, can be viewed successfully. However, systems that employ a non-polarizing lateral shear interferometer with an arrangement based, for example, on the Mach-Zehnder principle, produce successful and interpretable DIC images of both isotropic and anisotropic objects. This type of interferometer is discussed in Chapter 6.

5.5 ADJUSTING THE CONTRAST SYSTEMS

The important variable of an object for detection by MCS and DIC is the optical gradient. In transmitted light, the optical gradient is directly proportional to the product of the geometric gradient and the difference in refractive index: <p = a(nJ - n2)' In reflected light the difference in refractive index is effectively eliminated, which leads to <p = 2a. These variables in a field can have a wide range of values, thereby requiring each contrast system to provide a way of responding to these variations.

168 R. Hoffman

If the optical gradient is large the contrast may be too great. In the MCS, varying the effective slit width varies both the contrast and the relative coherence oflight impinging on the object. The contrast sensitivity can be reduced if the effective slit width can be increased. This is accomplished by allowing light to pass P2 by uncrossing PI with P2 (see Fig. 5.4). The effects of this can be analyzed from Fig. 5.6; by considering that contrast, C = (Bi - Bb)/Bb, where Bi is the brightness of the image and Bb is the brightness of the background. Increasing the slit width increases the intensity of the background. Changing the effective slit width also changes the coherence of the light at the object plane. The smaller the slit width the higher the spatial coherence anywhere in the object plane. The effect of excessive coherence is 'ringing' at the edges of the image of each object and a loss of edge definition.

The adjustment of the compensator in the DIC system provides for optimal images. Note that, when using Fig. 5.12, the intensity and/or color of the image will differ from the background according to the magnitude of the optical gradient. Let us say that the background is set for order ~ and the optical gradient is large, i.e. it creates a path difference of 0-400 11m (eqn (5.1)). Here the background will be grey (see Fig. 5.14) and one side will also be grey while the opposite side would be sky-blue with about the same intensity as the grey side. The image would not appear three-dimensional although opposite sides would have different colors. If the object were spherical and this large gradient was at the equator, there would be a continuous change of intensity from the center to the edge, with a dark region somewhere on one side of the object but not at the edge. The continuous change of intensity and/or color follows the intensity curve and 'abscissa' path difference from the background setting to the value of the largest path difference caused by the optical gradient at the edge. If these observations were made with a 20X objective whose shear distance d is twice as large as that of a 40X objective, changing to a 40x objective would reduce the maximum OPD to 0·200 11m, thus considerably changing the intensity distribution across the image. Generally, when the OPD (eqn (5.1)) is less than order ~, i.e. less than about 0·200 11m, the compensator can add or subtract a path difference such that one side is bright while its opposite is dark, and then the image appears three-dimensional. As the OPD increases beyond this value it becomes increasingly more difficult to create an intensity distribution across the image where the intensity varies from dark to grey to bright continuously across a sphere-like object.

Modulation Contrast and Differentiallnteiference Contrast 169

5.6 COMPARISON WITH THE PHASE CONTRAST MICROSCOPE

The phase contrast microscope outlined in Chapter 2 also enables phase objects to be viewed. However, the parts of the object that are detected are different from those in MCS and Ole. In phase contrast the sites of diffraction are converted to either a higher or lower intensity than the background.6 The primary sites of diffraction are edges and small particles. The greater the difference in refractive index the stronger the diffraction. Since the phase ring and light annulus in the phase contrast microscope are symmetrical, the detection in the system is symmetrical around the optical axis. The phase contrast system requires that the optical path through the object be small, i.e. less than one-tenth of a wavelength (often the case with polymer specimens), and that the object be flat. under these conditions the change in light intensity is fairly linear with respect to changes in the refractive index difference.

There are alternative objectives available, with different phase plates, to optimize the images of objects that do not conform to the above criteria.

The presence of a bright halo at the edges arises as an artifact from the system but can also arise in response to the optical gradient near the edges. As a result of either or both of these causes, the edges of an object when viewed with phase contrast are not reliably defined and it is a major advantage of the MCS and DIC systems that such halos are avoided. The halo effect in phase contrast is minimized as the refractive index difference gets very small and/or the feature becomes flat. Compared with MCS and DIC, the phase contrast image appears flat and is not affected by object birefringence. Any scattering feature will show contrast even though the sites may be closely packed and not resolved as discrete particles.

5.7 REFLECTED LIGHT WORK

The reflected light microscope is similar to a standard transmission microscope except that the illumination arises from above the objective. In this configuration the objective serves as a condenser when the light travels from the source to the object, and as an objective when light is reflected from the object. Reflected light is used routinely for examining

170 R. Hoffman

opaque specimens, such as moulding surfaces, but it can also be used to examine surface structures of transparent objects. As discussed in Chapter 6, the visibility of the object is dependent on the difference in refractive index between the object and its surround. The greater the difference the greater the visibility. However, whenever reflected light is used, any surface that the light meets on its way to the object will also reflect light back to the image plane. Therefore the surface to be examined should, if possible, be the first surface between the bulk of the object and the objective. Failing this, the object of interest should have a much higher index of refraction than whatever is interposed between the object and the objective. Light reflected back to the image plane that does not partake in image formation produces a loss of image contrast.

When examining an opaque object that has been coated with a transparent polymer, there are reflections from both the substrate/coating interface and the coating/air interface. Suppose that a highly reflective metallic surface is coated with a transparent polymer such as a polyurethane lacquer. To examine the topography of the metallic surface, the illumination and imaging beams must pass through the polymer coating. The polymer layer may be birefringent but the modulation contrast image is not affected by the presence of the polymer coating, whereas with DIC the detection of optical gradients on metal surfaces would be degraded by the presence of the polymer.

If we have a specularly reflecting metal surface then the internal microstructure of a transparent object on the metal surface can usefully be examined with reflected light.

5.8 IMAGE INTERPRETATION

We have seen that the image produced by MCS and DIC appears threedimensional, but this perception is subject to psychological artifacts, in that the image looks sometimes as an elevation and sometimes as a depression. These changes in perception can occur on merely blinking the eyes or viewing the same image at different times without changing any setting on the microscope. Also, different persons may see the image differently. This problem is well known as the 'depth reversal phenomenon'. In spite of this we tend to believe what we 'see'. However, the correct object contour can be verified by several methods. First, in


reflected light a known reference object can be placed in the field. Observe the shading when the shape is known. Secondly, in transmitted light, since the optical gradient is the product of two terms, to deduce the geometric gradient the relative refractive index must be known.

When using MCS, the object contour can be determined by examining the shading of the image using Table 5.1. Here the image of the slope at R is R' and the directions of the regions of the modulator are shown. If the relative refractive index is known (N} - N 2) then a rising slope, R (in this diagram looking at the object from left to right since that is the direction of the regions of the modulator starting with the dark region on the left), is bright at the image plane. Note the position reversal between the object and its image. In the first, fifth and sixth columns of Table 5.1, the optical gradient cp is shown. With the orientation of the modulator and the specimen as shown, a clockwise rotation of the deflection angle cp will cause light to pass through the bright region of the modulator.

When using DIC, positioning the compensator to the right or left of center reverses the shading. Therefore, to evaluate a known specimen, the position of the compensator should be indexed relative to its center position. In reflected light, the angle of deflection (optical gradient) is equal to twice the angle of incidence (geometric slope). The perception of an image as very curved or flat can also be an artifact. Changing the amount and the rate of change of intensity across an image can make a perfectly spherical object appear almost flat. These changes can be brought about by changing the difference in refractive index and/or the effective slit width in MCS or the effective shear distance in Ole. In MCS the effective slit width increases as the polarizer in front of the slit and the polarizer in the slit are uncrossed. In DIC, the shear distance decreases as the magnification increases. Owing to these variables the perception of curvature is open to misinterpretation.

The MCS and DIC produce images that are shaded in one direction. This is the direction where the object slope is most sensitive to detection. Ifwe consider an angle about the optical axis, and in a plane normal to the optical axis the sensitivity of slope detection changes as the cosine of that angle. If the objects of interest are generally round, it may not be important to rotate the specimen to detect all the slope directions; on the other hand, if the specimen is longitudinal, the object may be poorly detected if the slopes of the object are at right angles to the direction of maximum sensitivity.

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5.9 APPLICATIONS TO POLYMERS

The application ofMCS and DIC are generally qualitative and relate to the physical morphology and chemical nature (and hence refractive properties). These instruments can also be used as quantitative systems, but not easily. In reflected light, the optical gradient is merely twice the boundary gradient and represents a true geometric gradient. In DIC, the color changes rapidly about the first-order purple ('sensitive tint') background setting which is a sensitive position to detect small changes in the optical path difference. However, even though color recognition is subject to error, the slope can be approximated through eqn (5.1). Photometrically, the intensity of small regions of a slope of an object can be measured, scaled and applied to the known sin2 function using monochromatic light. The damped sin2 function, for the white light intensity distribution in Fig. 5.12, is replaced by a non-damped sin2

function when using monochromatic light. The data obtained are used to compute the optical path difference for that portion of the slope. If all the slopes on one side of the object are determined, then by integration the height can also be found.7 Using similar photometric methods, the slopes can be quantified with MCS using Fig. 5.6 and S = ftan </>, where S is the distance the slit image is shifted in the back focal plane of the objective and f is the focal length of the objective.8 However, quantitative investigations are best carried out using the techniques described in Chapter 6.

5.9.1 Surface Microscopy The characteristics ofMCS and DIC images make for the extensive use of these techniques in the study of polymer surfaces. A typical application is the examination of film surfaces. Films are often ideal specimens in that they require very little specimen preparation and can reveal a wealth of useful information which can be related to in-service performance and production variables. This theme is developed at greater length in Chapter 6.

A typical low density polyethylene film surface is shown in Fig. 5.16. Various levels of structure are well displayed by the MeS. The finest scale structure results from crystallization of the polymer. The larger structures, some 10 .um across, represent melt elasticity effects, whilst the largest visible features are due to extrusion die imperfections and surface damage.

By contrast, the very different surface of a polyester film containing

174 R. Hoffman

FIG. 5.16. Reflected MCS. Low density polyethylene film, showing surface structure due to crystallization of the polymer and melt flow effects. MCS

objective 4OX; reflected light. (X500.)

an anti-blocking additive is shown in Fig. 5.17. Here the surface structures are quite well separated on an almost textureless background. A third type of film, in this case film produced by biaxially drawing a propylene/ethylene copolymer, is shown in Fig. 5.1S. Both surface and sub-surface ring structures are visible; these are characteristic of the material and the process used for manufacture.

The sensitivity of both the MCS and the DIC system to surface roughness is remarkably high. Although the features seen on these films appear to be obvious, with considerable apparent depth, they are quite shallow structures (with the exception ofthe damage on the polyethylene film). In most instances this sensitivity is a boon to the microscopist concerned with investigations into the surface characteristics of films. However, the presentation of such images to film manufacturers or users is often followed by disbelief that the surface is so rough! The gross exaggeration of surface topography offered by these techniques can often give the impression that a given surface is 'unacceptably' rough,


FIG. 5.17. Reflected DIe. Surface of Melinex polyester film, showing surface roughness produced by anti-blocking additive in the film. Metallized (gold, sputter-coated) to increase contrast and mask effects from the highly

birefringent film. (X250.)

whereas in reality the surface is adequately smooth. This can be particularly disturbing when examining the surface finish of molds and casting drums. Even the most careful surface finishing techniques leave surface features that can be rendered visible by these contrastenhancing methods. As an example, Fig. 5.19 shows the surface of an acrylic replica (see Chapter 1) of a casting drum used in the manufacture of poly(ethylene terephthalate) sheet; cracking of the drum surface is clearly evident.

The role of these systems in relation to scanning electron microscopy is often queried since the resolution ranges overlap. In practice, provided the structures of interest are within the resolution range of the light instrument, DIC and the MCS can offer better image contrast and vertical sensitivity. Comparison experiments show that the visibility of shallow structures can be substantially greater even when compared with electron micrographs taken with a high specimen tilt angle, but this is true only if the surfaces being examined are relatively smooth. If rough surfaces are to be examined, scanning electron microscopy is the

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178 R. Hoffman

natural choice. Fracture surfaces are a good example; very smooth surfaces arising from low speed brittle fractures are frequently best examined by DIC or the MCS, but these techniques are most unsuitable for rough brittle fractures or where there has been ductility of the polymer.

5.9.2 Transmitted Light Applications As pointed out in Section 5.6, the DIC system and the MCS find the same areas of application as conventional phase contrast microscopy, discussed in Chapter 2. It is frequently the case that the refractive index fluctuations in a polymeric specimen are very small. If these fluctuations are due to the crystalline texture of the specimen, polarized light methods such as those described in Chapter 4 are often the most appropriate. However, this is not always the case, and phase contrast microscopy can sometimes yield superior results. The use of the MCS and the DIC system on such anisotropic specimens has already been discussed. Refractive index fluctuations will also arise in polymer systems in which more than one phase is present. A typical example of this is shown in Fig. 5.20 which shows a thin section of a polyester/ polycarbonate blend imaged using the MCS; phase separation is clearly

FIG. 5.20. Transmitted MCS. Thin section of polyester/poly carbonate blend in Cargille liquid of refractive index 1· 560, showing phase separation. (X 1000.)


visible. In another material (a 'high impact' grade of polystyrene) the rubber 'droplets' are clearly imaged using DIC (Fig. 5.21). It is interesting to note the elongation of these droplets which, incidentally, impart some anisotropy to the mechanical properties of the product. Indeed the impact strength of such materials as these is dependent upon the size of the droplets; monitoring of this by use of the techniques discussed in this chapter is therefore a routine part of diagnosis when mechanical failures are encountered.

Figure 5.22 shows a blend of Nylon 6.6 and polypropylene, imaged using a DIC system. Of particular interest is the local phase inversion which is clearly imaged in a dramatic way. It is tempting to see the image as having a three-dimensional character with raised or lowered areas, but this cannot be the real situation since we are here dealing with a parallel sided section (5,um thick). There is no interpretational difficulty provided that we are aware of the principles behind the formation of this type of image. Which phase looks raised or depressed is irrelevant; what the image is really telling us is the scale and distribution of refractive index gradients and fluctuations in the speCImen.

A particularly valuable application of a similar nature is the study of

FIG. 5.21. Transmitted Ole. Thin section of molding of high impact polystyrene. Note the elongation of the rubber droplets due to polymer flow effects

during molding. (X lOOO.)

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certain block copolymers. These may produce localized volumes in which the chemical composition is different from that of the surrounding matrix. The size and spatial distribution of these 'domains' will usually be influenced by the processing history of the material. As an example, Fig. 5.23 shows a DIC image of the microstructure of a propylene/ ethylene block copolymer similar to the type used to produce the film of which the surface is illustrated in Fig. 5.l8. The small (ethylene rich) 'droplets' exist within the spherulitic crystalline structure of the bulk polymer. It could be argued that here the DIC technique is being incorrectly used on an anisotropic specimen since the spherulites themselves are quite highly birefringent. It is true that interpretation of the morphology of the spherulitic structure itself could be difficult from this type of image, and modulation contrast might be more appropriate. However, here we are perhaps concerned less with the spherulites themselves, and more with the refractive index fluctuations produced by the domain structure. For this purpose the DIC technique is certainly working quite well. The contrast developed is satisfactory and there is no sacrifice oflateral resolution, as shown by the texture visible within the droplets which is on the scale of 004 Jim.

These techniques are not only applicable to polymer blends, composites or block copolymers. There are often instances where chemically homogeneous specimens need to be embedded or encapsulated in order to preserve their original morphology during thin sectioning. If the refractive indices of the specimens and the embedding materials are close, we again have a situation where the contrast enhancement techniques can be used to advantage. Such a situation is shown in Fig. 5.24. Low conversion poly(vinyl chloride) particles have been embedded in an epoxy resin. The peripheral membrane and primary particle structure are clearly visible when contrast enhancement is used; without the use of the MCS or DIC system the image information content is much reduced.

REFERENCES

l. Hoffman, R., The modulation contrast microscope: principles and performance,J Microscopy, 110 (3) (1977) 205.

2. Born, M. & Wolf, E., Principles of Optics. Pergamon Press, Oxford, 1975, p. 505. 3. Zemike, F., The concept of the degree of coherence and its application to

optical problems. Physica, 5 (1938) 785.

184 R. Hoffman

4. Padnos, M., Differentiation ofB and T lymphocytes in cell suspensions and smears. Nature, 259 (1976) 218.

5. Francon, M., Progress in Microscopy. Row Peterson, Evanston, IL, 1961. 6. Bennett, A H., Jupnik, H., Osterberg, H. & Richards. O. W .. Phase Microscopy:

Principles and Applications. Wiley, New York. 1951. 7. Croce. P., Laval, A & Sergent, B.. Integrateur eJectronique applique a

l'interferometrie. Revue Optique, 35(6) (1956) 359. 8. Hoffman. R. & Gross, L.. Modulation contrast microscope. Applied Optics,

14(5) (1975) 1169.

6

Interference Microscopy of Polymers

D. A. HEMSLEY Polymer Microscopy Services, Loughborough, UK

6.1 INTRODUCTION

There is a sense in which every light microscope can be seen as an interference microscope. This view has been discussed by Hartleyl who makes a clear distinction between 'conventional' and interference instruments on the basis of whether or not it is possible to adjust the phase difference between interfering beams in a controlled manner. This distinction is valid both in theoretical and practical terms. It is the availability of control over interference effects that allows true interference microscopy to be quantitative. This quantitative aspect is important, more so than the generation of image contrast, in that the information gained on polymer and polymer product microstructure can be of considerable practical significance.

Interference microscopy falls neatly in two parts in terms of historical development, equipment design and fields of application. On the one hand there are reflected light methods used for the study of surfaces. These have largely been developed with metallography in mind and have been widely used in that field. There is renewed interest in the technique for the monitoring of electronic components. As a result the necessary instrumentation is readily available commercially and the would-be polymer surface microscopist enjoys a wide choice of equipment. As we shall see, the final choice will in practice depend on the performance required, ease of operation and cost.

The second category of interference microscope of interest to the polymer microscopist is the transmitted light instrument. This has, in general; been designed with biological applications in view rather than

185

186 D. A. Hemsley

for the materials scientist or technologist. Indeed, polymer microscopists are in an interesting situation; although major potential users of both transmitted and reflected light methods, in all probability neither system was designed with their own applications in mind. This has the consequence that the polymer microscopist may need a combination of methods which the instrument designer did not envisage on a single microscope stand.

As with reflected light systems, the transmitted light methods discussed in this chapter are those used for quantitative measurement rather than image contrast generation. The latter requirement is usually better met by phase contrast, dark field, modulation contrast, differential interference contrast or polarized light methods which are described in other chapters.

In many ways synthetic polymers are ideal specimens for interference microscopy. In reflected light, surface structures of a height suitable for measurement by microinterferometry are plentiful. In transmitted light, the transparency of most polymers, at least in thin section, allows the ready measurement of refractive index, birefringence or geometrical thickness of phases present in the polymer specimen. We can conveniently interpret 'phases' here as meaning anything giving rise to localized refractive index fluctuations. Thus our definition will include effects due to crystalline texture and additives as well as phase separation in composite materials. The last type of specimen can be liberally interpreted to include, for example, the layers of different chemical composition in a laminated or coextruded multilayer film, provided that, by thin sectioning, this is observed sideways on. Possible applications for both transmitted and reflected light interference techniques are examined in more details later in this chapter.

Unfortunately interference microscopy appears to have gained a reputation for being in some way difficult to apply and producing results that are difficult to interpret. It is true that the setting up and operation of interference instruments requires attention to detail and an understanding of the basic principles. However, such an approach may be applied with advantage to all branches of microscopy and should not be seen as imposing special difficulties.

6.2 THE BASIC PRINCIPLES OF QUANTITATIVE MICROINTERFEROMETRY

In considering interferometry and interference microscopy we are intimately concerned with the wave nature of light which has been

Interference Microscopy of Polymers 187

discussed in generations of standard optical texts. That by Hecht and Zajac2 will provide a more than adequate modern treatment of the subject for those wishing to pursue a more detailed and theoretical treatment of interferometry. Consider the characteristics of the two waves depicted in Fig. 6.1(a), and note the inadequacy of representing a transverse electromagnetic vibration (a light wave) in this simple way; to describe a wave of this type more fully we would need to portray not only its amplitude (A) and wavelength (I\,) but also the direction of propagation, the orientation of the vibrations relative to some system of coordinates and the variation of all of these with time. Figure 6.I(a) shows only the first two of these parameters and their lack of variation over a period of only a few cycles.

Although the two waves shown have identical wavelengths and amplitudes, one lags behind the other along the time axis. The two

D

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waves E] and E2•

188 D. A. Hemsley

waves are said to be 'out of phase', and the phase difference is shown as cpo Because the wave motion is considered to be simple harmonic and represented by a sine function, we can conveniently use angular measure - degrees or radians. If our waves had started from the same point at the same time, the existence of cp implies that they must have followed different paths or travelled with different speeds.

In interferometry we must distinguish between the geometrical path length t between two points and the optical path length nt where n is the refractive index of the medium between the two points. Furthermore, when a wave passes through a series of media, 1,2, ... ,k, we can express the total optical path length as

x=k

x=1

The optical path difference (OPD) between two waves is an extremely important concept in interferometry and is related to the phase difference cp by the expression

271 cp = -(OPD)

A.

where A. is their common wavelength. This simple linear relationship between OPD and cp can easily be understood if we recall that one wavelength is represented by an angle of 271 in the generation of a sine wave. It is then clear that the fraction CP/271 is equivalent to OPD/A.. Equating these two fractions and rearranging gives the above expression.

We may reasonably ask how long the two waves shown in Fig. 6.1(a) will stay 'in step' and thus have a constant phase difference. No light source emits waves forming a continuous train, although for practical purposes a laser may usually be considered as doing this. In the case of the usual 'thermal' sources such as tungsten filament lamps and discharge lamps used in microscopy, waves are emitted as short trains of finite length. They are distinguishable from one another by abrupt and random variations in both phase and plane of polarization. Thus, if two waves originated from different sources, or different areas of the same source, we would not expect them to stay in step for very long. If, however, the two waves are derived in some way from precisely the same part of the same source, we would expect cp to remain constant, since any change in the emission will affect the two waves equally.

The above discussion introduces the idea of wave coherence. It is


convenient here to distinguish between two aspects of coherence, although at the deeper levels of theory such a distinction disappears. Consider a point or infinitely small source of light emitting waves passing along any particular radius from the source. Suppose we now compare the phase difference cp between two chosen points P I and P 2 on that radius. Clearly, if the wave trains from the source are continuous for a time large compared with the time for the wave to move from PI to Pb

the phase difference will generally be constant. Increasing the distance between PI and P2 will eventually make it improbable that a constantcp can be measured. In this case the distance from PI to P2 (d!. 2) exceeds the 'coherence length' of the radiation from the source. Alternatively, we might think in terms of the 'coherence time' which is given by d1,2/c where c is the velocity of light. The coherence time is closely related to the degree to which light can be considered to be of a single wavelength, i.e. monochromatic. Strictly only a wave showing infinite coherence time (clearly a practical impossibility) would be truly monochromatic. Fortunately we are normally content to work with 'quasi-monochromatic' waves having finite bandwidths and with lower, but still satisfactorily large, coherence times.

The practical significance of the coherence length will be emphasized later when the limits of microinterferometric measurements are discussed. In the remainder of this text the term 'monochromatic' may be read as 'quasi-monochromatic' by those who prefer a more rigorous approach.

The second aspect of coherence we need to consider is the concept of 'spatial coherence'. Suppose PI and P2 are not now on the same radius from the point source, but represent two points some distance from an extended or multiple source. An examination of the phase difference between waves at PI and P2 might be expected to show no coherence, except for the fact that all of the source is emitting common wavelengths. However, it can be shown3 that in interferometry the angular size of the source has a marked effect on the visibility of interference phenomena. As the angular size increases, so the spatial coherence decreases, as does the visibility. Again, this has practical significance influencing the conditions of operation of interference microscopes.

A detailed exposition of coherence, which is inevitably mathematical, would be out of place here; readers are referred to Born and Wolf' and Francon3. 5 for a more detailed analysis.

Consider a model experiment in which two coherent waves pass

190 D. A. Hemsley

through the same point in space. In general there will be constant amplitude and phase difference cp between them, and these dictate the characteristics of the 'resultant' wave produced. Figure 6.1(b) shows how the resultant is derived by the addition of instantaneous amplitudes. If the amplitudes of the two original waves are equal (this is usually the case in practical microscope interference systems) then it is clear that for a phase difference of TT (or 180°) the result of the addition is constantly zero and 'destructive interference' has occurred. This will happen for any phase difference TT where p is an odd integer. Other phase differences, or differences in amplitude, will not permit a zero resultant to be achieved. Another special case arises when the phase difference is 0 or any multiple of2TT. The resultant then has a maximum amplitude.

In practice it is not the amplitude of the resultant wave but the observed irradiance that is important. This is given by the square of the amplitude. Thus, for waves of equal amplitude (A), the irradiance (1) will vary between zero and 4A 2 according to the phase difference cp between the waves.

Some further comments on this interference effect will be useful, before formally listing the conditions under which the phenomenon will occur. It should first be emphasized that the effect will be evident only if coherent waves are involved. Suppose we attempt to observe interference phenomena between waves emitted by two· independent thermal sources. Since the phase of each wave is changing after short periods of emission (of the order of 10-8 s) no persisting interference effect will be seen. It can easily be shown that the irradiance at a point will then be the algebraic sum of the irradiances from the two sources. It is therefore impossible to construct an instrument using the interference phenomenon, such as an interference microscope, by using more than one source. This point needs to be qualified if we consider the possible use oflasers which emit highly coherent light. It has been demonstrated that two lasers can be used to produce a stable interference effect although, to the best of the present author's knowledge, no interference microscope has been constructed using this approach.

Although for simplicity all of the above discussion has concentrated on the combination of just two waves, identical principles are involved for the combination of greater numbers; as described later, a commercially available microscope interferometer utilizes this fact.

The above discussion has also avoided consideration of the state of polarization of the waves involved; this is of considerable importance


from both the theoretical and the practical standpoints. Both Fresnel and Arago made a detailed study of the role that polarization plays in interference. A formal statement of their results can be found in the literature4 as the Fresnel-Arago laws; in essence they indicate that interference cannot occur between plane polarized waves that have their planes of vibration mutually at right angles, but if the two vibration planes are the same then interference will result. The practical importance of these laws is recognized when one sees that for transmitted light interference instruments a popular method of achieving two equal amplitude coherent beams is to split a plane polarized beam using doubly refracting optical components; the resulting two beams are then orthogonally polarized.

Taking into account the characteristics of coherent waves, we can now list the conditions for the production of optimum interference effects by stating that the interfering waves must have:

- the same wavelength; - a constant phase relationship; - the same plane of polarization; - identical amplitudes.

So far only the interference effect occurring at some chosen point in space has been considered, but in an interference microscope we are concerned with a field of finite dimensions. It is quite possible, and in some cases desirable, that the phase difference between interfering waves varies from one part of the field of the microscope to another. The conditions for destructive interference will be satisfied at some points but not at others. A pattern of field irradiance will then be observed which is termed an interference pattern. From what has been said it will be clear that the visibility of such a pattern, and disturbances of that pattern produced by a specimen, depends on the relative phase differences involved and the relative amplitudes. Fringe visibility, or contrast, which can be defined as

where Bm• x and Bmin are the maximum and minimum fringe irradiances, will be a maximum when Bmin = 0, i.e. the amplitudes of the destructively interfering waves are equal.

The quantitative use of interference microscopy relies on the

192 D. A. Hemsley

interpretation of interference patterns produced by specimens in terms of the phase shifts or, as we saw earlier, OPD.

In some cases the correctly adjusted interference system generates a set of interference fringes across the field of view of the microscope, and measurements are made by observations on the disturbance of this, usually linear, fringe pattern. In other systems the fringe spacing may be extended by adjustment so that it is effectively infinite, and compensation methods are used as described later.

It is now convenient and logical in terms of the application to polymers, and commercial availability, to consider reflected light and transmitted light systems separately. The underlying principles are nevertheless often very similar, or even identical.

The interference systems described below are often available as 'add on' extras for an existing microscope stand, from the same manufacturer. In a few instances units may be easily fitted to stands from a range of manufacturers. However, this is much more common with reflected light interference systems than with transmitted light systems.

A basic requirement for any interference microscope is mechanical stability; in practice this may preclude the fitting of interference optics to the less robust microscope stands. On the same theme it must be added that interference instruments need ideally to be operated in locations virtually free from vibration. The sensitivity of these instruments is such that, in modern buildings of relatively light construction, they can detect the slamming of a distant door or passing footsteps. These problems become particularly acute when carrying out photomicrography. The most sensitive instruments should also not be sited near heat sources which may produce convection currents giving density and thus refractive index fluctuations in the air in the region of the specimen.

6.3 REFLECTED LIGHT APPLICATIONS

6.3.1 Specimen Preparation The visibility of interference effects using reflected light methods is strongly influenced by the reflectivity of the surface under investigation. The examination of the surface of a polished metal would normally present no problems but polymer surfaces have relatively low reflectivity and image contrast can be poor under all but ideal conditions. Furthermore the transparency of polymers means that even in compounds containing pigments or internal fillers there is substantial


penetration oflight through the surface and into the material. Here the light will be scattered and a proportion will return through the surface and contribute to image formation. Surface image contrast can be noticeably reduced by this mechanism. Even in the case of highly transparent polymers in which the internal light scattering is negligible, reflected light from the second surface of the material may cause image degradation. This is particularly noticeable in the case of the thin highly transparent films which can be produced from polypropylene poly(ethylene terephthalate) or cellulose acetate.

Both the problem of low reflectivity and that of internal scattering can be eliminated by coating the surface of the polymer with a thin layer of metal, either by sputter coating or by evaporation coating (see Chapter 1). Both are well established techniques and are not discussed in detail here. However, it should be pointed out that either process can, unless care is taken, lead to excessive heating of the polymer surface with a consequent modification of the surface topography. With reasonable efforts to minimize heating, such as the provision of cooling, good thermal contact with the specimen being coated and the deflection of unwanted electrons, success is possible with all but the most sensitive specimens. For interferometry the choice of metal is not normally critical and the selection is made more on the basis of sputtering efficiency or evaporation characteristics than on the optical properties. Gold is traditionally used for sputter coating and aluminium for evaporation. An advantage of the use of gold in sputtering is that the same specimen can then, if necessary, be examined by scanning electron microscopy without much, if any, further preparation.

It is possible that metal coating facilities are unavailable, or that time is at a premium, as may be the case in a quality control laboratory. With thin specimens such as films, one is able to make a substantial improvement to the reflected light image by reducing the reflected intensity from the lower surface by the application of a light-absorbing layer. This is conveniently done with black ink from a felt-tipped pen! The result of applying this technique to a polyethylene film is shown in Fig. 6.2 which clearly shows increased contrast from the treated area.

The surface reflectivity of a polymer for light at normal incidence can be estimated from Fresnel's equations4 and a knowledge of the refractive index (n) of the polymer. Thus the percentage reflection R (in air) is given by

R= -- XlOO (n - 1)2 n + 1

FIG

. 6.2

. Im

age

(X 3

(0)

of t

he s

urfa

ce o

f pol

yeth

ylen

e fi

lm. T

he a

rea

A is

as

norm

ally

imag

ed. T

he a

rea

B h

as b

een

trea

ted

wit

h bl

ack

ink

on

the

rev

erse

sid

e to

inc

reas

e th

e co

ntra

st b

y re

duci

ng s

econ

d su

rfac

e re

flec

tion

s.

i t::::l

;>:.. ~ ~ ~

Inteiference Microscopy of Polymers 195

For polyethylene film n is approximately 1· 51 and for poly( ethylene terephthalate) film n = 1·62 (mean values). These refractive indices lead toR values of 4·1 and 5·6 respectively. This difference is quite noticeable in practice and allows the successful direct observation of poly( ethylene terephthalate ).

The examination of surface roughness may be indirect, through the use of a surface replica. Replica production has been discussed in Chapter 1, so it is sufficient here to reiterate the two important advantages of replication.

First, it is useful to replicate inconveniently large specimens which cannot be cut into a size suitable for coating or putting on to the microscope stage. In effect, replication makes the surface examination non-destructive. This might be particularly important, for example, in determining the surface roughness of a film casting drum or of a large and expensive mould or moulding. The second advantage is that successive replicas from the same area of the specimen can be used to show changes in surface roughness over a period of time. The same area is replicated repeatedly and the replicas are stored for subsequent examination and comparison.

6.3.2 Applications The application of reflected light differential interference methods to the study of surfaces is examined in Chapter 5. For qualitative work this, and modulation contrast, has proved to be a powerful system, greatly increasing the visibility of surface topography on a wide range of polymer products. Such applications have been extensively discussed in the literature, and there has been some discussion of the use of this method in quantitative applications. In the present chapter we are concerned more with techniques that allow surface topography to be quantified by the direct measurement of optical path differences.

Ofthe many products produced from plastics and rubber, films are of particular interest when considering surface topography. Many of the more important properties of films, such as transparency and frictional characteristics, depend heavily on the surface roughness.

Considering transparency, light scattering will generally occur both within the film and at the film/air interface. The latter is particularly intense because of the high refractive index difference between the polymer and the surrounding air.

The amount and angular distribution of the scattered light will be manifested visibly in a number of different ways. Thus the appearance

196 D. A. Hemsley

of a film may be described subjectively by terms such as 'haziness', 'clarity', or a variety of others - often poorly defined. Strict definitions can come only from consideration of the distribution of scattered intensity as a function of the angle of scatter, but it is easily shown that structures on the surface of a film that are of a size visible to the light microscope will more affect the ability to resolve detail viewed through the film than give an impression of haziness. An examination of the surface roughness of a film by quantitative interferometry might therefore be expected to yield information correlating with film clarity. Such proves to be the case.

The frictional characteristics of a film are, together with its stiffness, very important in determining 'handling' properties in such demanding processes as high speed packaging, as well as in controlling the 'feel' of the film in other less critical applications. Closely associated with film handling is the phenomenon of 'blocking'. The difficulty of separating two closely contacting smooth sheets of material such as glass microscope slides is widely experienced. Polymer films, particularly those manufactured from polypropylene and poly( ethylene terephthalate), can exhibit similar characteristics if their surfaces are especially smooth. In extreme cases it may become virtually impossible to separate layers of the film or even to unwind a roll of the material. To prevent such behaviour it is common practice to ensure that the surface has a controlled amount of roughness. This may be achieved by the inclusion of a particulate inorganic or polymeric filler which will generate small surface excrescences. In practice the size distribution, optical properties and concentration of filler must be carefully chosen since there is a conflict between the need to control blocking and friction through surface topography, and the production of excessive surface light scattering which destroys the clarity. Other sources of surface roughness on commercial film include coatings or surface defects arising from interactions between the polymer melt and the processing conditions. A wide variety of such defects can occur, often specific to a particular process. Indeed, qualitatively these may be used to identify a manufacturing process or even the film from a particular manufacture.

The effectiveness of particulate additives or other routes in the production of anti-blocking surface topography can be assessed using surface interferometry. Figure 6.3 shows the surface of a film viewed using this technique. The precise interpretation of such images is discussed later but, if at present the dark fringes are taken to represent surface contours with a vertical spacing of about 250 nm, some

FIG

. 6.3

. S

urfa

ce o

f a

poly

mer

film

(X

238)

im

aged

usi

ng a

sur

face

int

erfe

rom

etry

tec

hniq

ue.

:; ;;; ~

~

;:, ~ ~ <3 ~

~ '" ~ ::? ~

2! ~ §

198 D. A. Hemsley

appreciation can be gained of the type ofwark that may be carried out using the technique. Process-induced rough surfaces or anti-blocking additives play an important part in determining the frictional characteristics discussed above. However, it is in practice often necessary to include a 'slip additive' into the formulation of the film. These, usually wax-like, additives provide lubrication of the surface often as a thin layer. They are less easily detected by interferometry but the possibility should not be ruled out.

Other film or foil surfaces that can profitably be examined include those produced by post-extrusion processes such as embossing, coating or printing. The exact contours of an embossed area can reveal the depth and effectiveness of the embossing process and even the condition of the equipment used. Figure 6.4 shows the surface of an embossed poly(vinyl chloride) foil; clearly at the point examined there is saddle-like embossing.

The surface topography of coated films may arise from 'submerged' structures on the base film or from the coating layer itself. Local removal of the coating using a suitable solvent and comparing surface topography will clarify the situation. Measurement of the thickness of thin coatings laid down on film is also possible by simple interferometry if a reasonably steep step can be produced by the same process of solvation of the coating. Ifthe coating substrate adhesion' is poor then physical methods of removal may be attempted but care has to be exercised to avoid thinning of the coating adjacent to its edge when measurements are taken. In either case it is the interference fringe displacement occurring at a coating edge that is measured. Figure 6.5 shows the step due to the removal of a coating from a PVC card.

The surface topography of moulded products often replicates the roughness of the moulding tools. Even highly polished tools show some marks on the. microscopic scale. The faithfulness with which the polymer reflects these marks depends upon a number of variables such as the moulding pressure, the viscosity of the melt, mould design and whether crystallization of the polymer is involved. For a given material and mould, a comparison of the interference pattern exhibited by a test moulding can be compared with that given by a reference moulding, or the tool or (more conveniently) a prepared replica of the tool. The relative topography can then be related to moulding parameters. It is prudent to compare identical areas since the surface roughness may vary from place to place.

In the case of crystalline polymers, and where there is poor contact

FIG

. 6.4

. S

addl

e-li

ke e

mbo

ssed

pat

tern

on

the

sur

face

of

a PV

C f

oil

(X 1

05).

:: " ~ ~ ;:, R ~ <:l ~ ~ '" ~ cl' ~

~ 3 ~

FIG

. 6.

5.

A s

tep

in t

he s

urfa

ce o

f a

coat

ed P

VC

car

d. A

rea

A s

how

s th

e n

orm

al c

ard

sur

face

an

d a

rea

B sho~s w

here

a

coat

ing

laye

r h

as b

een

rem

oved

(X

113

).

N 8 !::l

;.. ~ ~ ~


with the mould surface or contact is totally absent, it is usual to observe spherulitic texture on the surface of the moulding (Fig. 6.6).

It has been assumed that the surface of the moulding is essentially smooth. Any intentional texturing of the surface, to produce a matt finish or simulated wood or leather grain, will usually inhibit or prevent measurements by interferometry because the surface is too rough.

Other typical applications of surface interferometry include the determination of the radii of curvature of plastic lenses, correlations between surface roughness and gloss, and the investigation of the severity of surface damage produced by wear and abrasion.

6.4 INTERPRETATION OF SURFACE INTERFEROGRAMS

As discussed in more detail later, the general construction of most reflected light interference microscopes allows for the following operations:

l. Production of a beam of either white or monochromatic light. 2. Splitting of this beam into two or more coherent beams. 3. Reflection of the beams by both a reference surface and the

surface of the specimen. 4. Recombination of the beams to give an interference pattern.

Consider the simplest possible experimental system shown in Fig. 6.7. Here we have two surfaces, the specimen surface S and the reference surface R; the latter is semi-transmitting and performs a double role as beam splitter and reference. Light from X is reflected at both surfaces and the optical path difference (P) between the two beams is given by

P = 2d cos () (6.1)

where d is the separation of the surfaces and () is the angle shown. Note that we have confined our attention to just two beams and have ignored possible further reflections of the beam leaving S at the underside of the surface R.

The condition for constructive interference ofthe two reflected beams at Y to give a bright fringe will occur when P = (n + !), where n is an integer; this takes into account the phase shift occurring at the specimen surface. Thus

n +! = 2d cos () (6.2)

FIG

. 6.

6.

Sph

erul

itic

cry

stal

liza

tion

str

uctu

res

on

the

sur

face

of

a po

lypr

opyl

ene

mou

ldin

g (X

338)

.

N o N ~

t>- ~ ::! ~


x

R

s-----.",..,.--..L

y

FIG. 6.7. A basic two-beam interference system.

Clearly, if d varies as a result oflarge height changes on the surface S, a series of bright and dark fringes will be seen with a 'contour' interval of M2. In the total absence of surface roughness (an unlikely situation with polymer surfaces), the field of view would be uniformly bright. In practice d may vary as a result of a wedge angle between surfaces Rand S giving a field covered by linear fringes. Roughness of the surface S would then superimpose a local modification on this regular fringe pattern.

Step heights on surfaces can be measured and may in turn be used to measure the thickness oflayers on, for example, coated films as already discussed. Provided that a clean edge to the coating is available, one simply measures the fringe shifts occurring across the edge. If the step is fairly broad it is possible to follow an individual bright or dark fringe from the base film to the surface of the coating. A sharp step however can present more of a problem. The use of monochromatic light does not allow individual fringes to be recognized and, although the fractional fringe shift across the step can be measured, the integral number of fringes remains in doubt. However, we can 'label' individual fringes by using white light. In particular, the black 'zero order' fringe can readily be used to establish the integral number of fringes to which the fractional fringe shift must be added before the step height is determined. Using the two-beam interference method it should be possible to determine step heights to better than M20 (25 nm for green light).

The above interpretation has neglected some complicating factors which must be recognized. In particular, we have assumed that cos () in eqn (6.1) is constant and unity, corresponding to near normal incidence. This amounts to the microscope system having a small numerical aperture (NA), which militates against obtaining adequate resolution of small surface features such as the surface bumps produced by antiblocking additives. A larger NA illuminates the specimen over a range

204 D. A. Hemsley

of angles from 0 to sin -I(NA) (assuming the specimen to be in air), and in practice a weighted and averaged cos () needs to be used in all calculations. Most important, from eqn (6.2)

d n +! __ 2_

2 cos ()

and the fringe spacing is clearly dependent upon the illuminating aperture. In practice a fixed but unknown value of the numerical aperture is used based on the choice of objective lens and instrumental settings such as the aperture iris. The system can then be calibrated using a standard specimen. This yields a mean value for cos () which can most conveniently be used in all calculations as a modifier of the wavelength used. Thus instead of using A we use A!COS7J.

The steepness of surface slope that can be measured is ultimately limited by the need for the reflected light to enter the aperture of the objective lens. In principle therefore high NA lenses can accommodate steeper slopes than low NA lenses. However, the situation is more complicated since, in acquiring a high NA, depth offield is sacrificed, so the top and bottom of surface features in the field of view cannot be imaged simultaneously. Excessively steep regions on the specimen give rise to dark featureless areas in the microscopic image (Fig. 6.3) and, even before this condition is reached, large wedge angles between reference and specimen surfaces can lead to significant measurement inaccuracies, so such angles are best avoided in practice.

Another complication is that the coherence length of the light used must be greater than P, otherwise interference fringes cannot be obtained. The practical significance of this fact is that it is usually much easier to set up an interference system with a high coherence source such as a low-pressure discharge lamp than with white light. A highpressure discharge lamp and a suitable interference filter to select a strong spectral line (such as the 435 nm or 546 nm lines in the mercury lamp spectrum) also provides satisfactory coherence for most purposes. The standard procedure for measurements that involve the use of both monochromatic and white light, such as the step height determination outline above, would involve initial adjustment using monochromatic light before recourse to white light fringes. Furthermore, since the coherence of a white light source is substantially reduced, very large height or step measurements may be impossible to measure.

InteJference Microscopy of Polymers 205

White light fringes may also assist in deciding whether, for example, the area Yon the embossed profile shown in Fig. 6.4 is higher or lower than the surroundings. Valleys or hills on the surface will give identical monochromatic light fringe patterns regardless of instrumental adjustment. On the other hand, the behaviour of a white light 'labelled' fringe can be examined as the position of the reference surface is moved towards and away from the specimen. Provided that the instrument has sufficiently fine control of this adjustment, and the direction of movement of the reference can be deduced, the problem is solved. An alternative, often simpler approach is to ensure that a purposely introduced feature such as a scratch is produced on the specimen to provide a reference against which other features may be assessed.

One of the main disadvantages of two-beam systems is that the fringe irradiance is a cos2 function of position, so the bright and dark fringes are broad and of equal thickness. It is rather similar to having thick contour lines drawn on a map. The elevation of a point falling within a contour line cannot precisely be determined. For greater measurement precision we need thinner contours on the map. In interferometric terms these can be achieved by using a multiple-beam rather than a twobeam interferometer. The method is briefly described below and at much greater length in the literature.6 The interpretation, as opposed to the application of the method, is simple. A typical multiple beam interferogram is shown in Fig. 6.8. In order to obtain a multiple beam image and to satisfy coherence requirements, the illuminating cone angles must inevitably be small, so that the fringe interval can conveniently be taken as exactly Al2. As can be seen, fringe widths are very substantially reduced and the improvement is such as to allow the measurement of fringe displacements of around Al200 (2, 7 nm for green light). This method is most frequently used for the comparatively simple task of step height measurement.

A final and essentially practical point on interpretation is that this is usually best done from micrographs rather than directly from the image in the microscope. Apart from overcoming such problems as vibration effects and low light intensities, the processing of micrographs provides an opportunity for enhancing fringe contrast. Indeed, photographic processes exist (e.g. Agfacontour) for selecting and printing a particular grey level from this image, and this can be used to increase the precision of measurement of cos2 fringes. Electronic methods of image enhancement are also available; these may be used alongside standard image analysis techniques to determine fringe spacing.

FIG

. 6.

8.

A t

ypic

al m

ulti

ple-

beam

int

erfe

renc

e im

age

of

a p

oly

mer

fil

m s

urfa

ce (

X 1

05).

~ I:::l

:.:.. ~ ;:: "' ,~


6.5 SOME REFLECTED LIGHT SYSTEMS APPLICABLE TO POLYMERS

6.5.1 The Coverslip Method

207

A very simple but often useful 'interference microscope' can be constructed with the aid of a flat coverslip. This is coated with a thin semi-reflecting layer of a metal such as gold (by sputtering) or aluminium (by evaporation). The thickness of the coating is not particularly critical and the optimum will vary according to a number of parameters, including the reflectivity of the polymer surface under investigation. A coverslip reflectivity of about 50% is acceptable for most purposes but some experimentation with higher or lower values may produce superior results. Ideally the polymer surface should also be metallized to increase its reflectivity.

Any microscope equipped for normal bright field reflected light work can be utilised, provided that there are facilities, usually in the form of an aperture controlling iris, for restricting the angle of the cone of rays falling on the specimen. Relatively low-power objective lenses (say X 10 or X20) are used. A reasonably high intensity, high coherence light source is required, and typically this will be a low pressure sodium lamp or a high pressure mercury arc lamp with a narrow band interference filter.

The coverslip is simply placed on the area of the surface of interest with its metallized surface in contact with the polymer. Interference fringes which are essentially two-beam in character will be seen giving the contours of the surface. Obviously this simple system does not provide for adjustment of fringe spacing or variation of the referencespecimen separation. Nevertheless the image is usually adequately interpretable and, in the absence of more sophisticated equipment, provides quantitative surface information at virtually zero cost. The interference fringes are increased in visibility by restricting illumination of the surface to a narrow cone of rays. Closing down the aperture iris to achieve this degrades lateral resolution but increases fringe contrast, so a subjective compromise must be sought that is appropriate for the particular specimen being examined.

This compromise has to be made in many types of microscope interferometer. Another factor which might be expected to degrade resolution is the use of a reflected light objective lens in conjunction with a coverslip. Normally these lenses are designed to be used without a coverslip, but at low magnifications, and bearing in mind other factors

208 D. A. Hemsley

militating against high resolution, the extra deleterious effect of the coverslip is usually negligible.

6.5.2 The Watson Type Microscope Interferometer This device is an attachment which may be fitted to almost any microscope, whether or not designed for reflected light work, and replaces a normal objective lens. The principle of operation is essentially that of a Michelson interferomete~ and is shown in Fig. 6.9. The source (L) may be a low pressure sodium lamp, a high pressure mercury lamp or a tungsten filament lamp for white light work. Light from this source passes down the horizontal tube, containing a filter if required, to a beam splitting block (B). Half the light is then reflected down on to the specimen surface (S) and half passes through the block on to a front silvered reference surface (R). On returning from the reference and the specimen, the reflected light is again split by the block to give light passing back towards L or upwards to the microscope objective lens (0). Two sets of beams therefore contribute to image formation by the microscope, one coming from the specimen and the other from the optically flat reference surface. The block to reference distance and the angle of tilt of the reference surface are adjustable by means of screw controls. Optically the reference surface may therefore be brought into near coincidence with the specimen surface, and interference between the two beams entering the objective lens can readily be obtained. Control over the spacing and direction of the interference fringes is by the reference tilt adjustments. The necessary conditions for coherence are satisfied in general since the two beams contributing to the final image will have originated from identical parts of the light source and have travelled almost equal distances. The latter requirement is not met

s

FIG. 6.9. The basis of the Watson type of microscope interferometer.


if the adjustment of the block to reference surface distance is shorter or longer than the block to specimen distance by more than the coherence length of the light. For this reason it is advisable to use a high coherence source such as a low pressure sodium lamp, at least during initial setting-up. Attempts to set up the equipment from scratch using a white light source are a prime cause of frustration with this type of interferometer. If white light fringes are required, good fringes in the desired orientation should first be obtained using a highly coherent source before replacing this with a filament lamp and carrying out fine tuning.

This commercially available device is relatively inexpensive and has much to recommend it for many polymer applications. It is available with either 8 mm or 16 mm objective lenses, giving primary image magnifications of X20 or X 10 at a standard 160 mm tube length. However, two shortcomings are evident in use. In certain applications, such as the observation of very small surface bumps on films, or surface spherulitic crystallization on mouldings, or fine wear scratches, the adequacy of the lateral resolution of the system is doubtful. Inevitably the placing of the beam splitting block in front of the objective lens limits the NA of the system, which in turn imposes severe limits on resolution. This is of little consequence in the observation and measurement of step heights occurring at the edge of gross features such as surface films or etch pits, but it is more important in the cases mentioned above.

The second problem is one of mechanical stability. The relatively simple design results in the equipment being prone to vibration problems unless precautions are taken as to where and how it is used. Nevertheless this equipment allows measurement to the usual precision of a two-beam interferometer, although this is best done from photomicrographs.

It is perhaps finally worth noting that it has been reported7 that this device can be used in unison with a differential interference contrast system to give simultaneous quantitative and qualitative information.

6.5.3 The Linnik System Many of the problems associated with the Watson interferometer are overcome in the Linnik system, although the basic principle of operation is essentially similar as shown in Fig. 6.10. Comparison with Fig. 6.9 shows that both light paths in this two-beam system now contain ·objective lenses, and beam splitting and recombination takes place behind rather than in front of the lens imaging the specimen.

210 D. A. Hemsley

B R

~--~~+-----~~-----------x L

s_ ......... _

FIG. 6.10. The basis of the Linnik microscope interferometer.

Consequently there is less constraint on the usable objective aperture and image resolution is substantially improved. Matched pairs of objectives are available covering the normal range of primary image magnifications. The 'reference' objective is focused on a reference surface the reflectivity of which may conveniently be chosen to match that of the specimen and thus optimize contrast. This feature can be of special value when examining the surface of uncoated polymer. The Linnik system lends itself well to the provision of adjustment to control fringe orientation, spacing and the equalization of optical paths. In its commercial form it is a robust unit designed to eliminate virtually all vibration problems. In use the advantages over the Watson system are obvious but they are obtained at a substantial increase in cost. Nevertheless, for routine use on polymer surfaces the Linnik system has much to recommend it.

6.5.4 The Mirau System This two-beam system, the principle of which is illustrated in Fig. 6.11, differs from those already described in that both the beam splitter (B) and the reference surface (R) are positioned between the microscope objective (0) and the specimen (S). However, both are sufficiently thin not to increase the working distance excessively and hence reduce the NA of the objective lens. Objective magnifications up to x40 are available commerically.

An easily selected range of reference surface reflectivities is normally

From light source

Q) >- c ~ ., ., -E 0. .- Q) ~ Cl c.., ~ .5


M

L+-----I-' 0

FIG. 6.1l. The basis of the Mirau microscope interferometer.

available so that fringe contrast is readily optimized. The potential for mechanical stability in the design is satisfactory and adjustment of fringe spacing and orientation is again achieved by tilting the reference surface.

This system has found wide use in the field of plastics films and is sufficiently robust and inexpensive for it to be used in quality control laboratories for routine monitoring of film production.

6.5.5 The Reflected Light MUltiple Beam System A number of microscope manufacturers have reflected light multiple beam interferometers in their catalogues; the principle is illustrated in Fig. 6.12. They are sometimes referred to as Tolansky systems following the extensive development and application work carried out and reported by Tolansky.6

As discussed above, multiple beam interferometry gives narrower fringes and allows substantially improved vertical resolution. In practice, and with commercially available systems, the conditions of operation are not normally such that the full potential of the multiple beam method can be realized. Nevertheless significant improvements

212 D. A. Hemsley

FIG. 6.12. The principle of the multiplebeam interferometer. S represents the surface of the specimen and R a reference surface having a high reflectivity. Light entering the system from 0 suffers successive reflections at Sand R. At the latter surface some light emerges (waves W h W2_ W3 etc.) and is used in

image formation.

o

---\-~r-Ir-+-+--+---- R

over two-beam methods are possible and the system is recommended where maximum precision of height measurement is required. Typical applications would include thin film thickness measurement and the examination of surface roughness produced by the incorporation of small particulate additives.

The system hardware usually consists of a special objective assembly which replaces a normal bright field reflected light objective. Also supplied are a set of semi-reflecting 'reference flats' which are mounted in front of the objective in contact with (or extremely close to) the surface under examination. It is usual for a range of reflectivities to be provided so that these may be matched to the specimen. Typically this range is 4-80% reflectance. For special applications the reference surfaces may themselves be curved, e.g. cylindrical, to allow the routine observation of slightly curved specimen surfaces.

Fringe spacing and orientation are achieved by slight tilting of the reference surface, and a control giving vertical axis movement ensures maximum fringe visibility when the specimen is in focus.

The main disadvantage of the simpler systems is mechanical stability - they are unsurpassed as laboratory seismometers! A second problem is that the semi-reflecting metal layer on the reference surface may become damaged by contact with the specimen surface, particularly if the latter is rough. The specimen should also not be moved without first raising the reference surface to provide clearance.

6.6 TRANSMITTED LIGHT INTERFERENCE MICROSCOPY

6.6.1 General Comments on Transmitted Light Measurement With a transmitted light interference system we have the ability visually to assess or accurately measure optical path differences (OPD) between waves passing through a chosen area ofthe specimen and those passing


through a reference region. Usually this reference region is within the field of view of the microscope but its precise location varies according to instrument design.

Assessment ofOPD may be carried out visually by the observation of interference colours (in white light) or interference fringes (in white or monochromatic light). Accurate measurements ofOPD are made using 'compensator' systems; however, the visual assessment is always a useful and rapid check that sensible compensator readings have been obtained. The use of compensators is discussed in more detail below. It is sufficient here to indicate that generally they are 'null' devices in which the OPD to be measured is reduced to zero by the compensator providing an equal OPD but in an opposite sense.

As shown in Section 6.2, the OPD is a function of both refractive index and specimen thickness. Obviously, given one of these, the other is readily obtained from OPD measurements. Many situations occur in which neither quantity is known; in these cases either a double experiment is necessary in which the refractive index of the reference region is changed, or use is made of special geometrical considerations such as the specimen being spherical (e.g. latex particles) or cylindrical (e.g. fibres).

The most common reasons for refractive index measurement are specimen identification, determination of chemical composition or molecular orientation determination. Some examples of applications specific to synthetic polymers are given below and are chosen to illustrate the wide application of this powerful microscopical method. However, it should be remembered that, as for reflected light microscopes, the lateral resolving power of the microscope is at best unchanged by the incorporation and operation of the interference equipment and in practice may be substantially reduced. It is therefore unusual for microinterferometry to be carried out on image features less than 2 pm in lateral extent, and even this is beyond the scope of some types of interferometric equipment. The limitation on applications that this resolution limit imposes is in practice not especially severe since many important structural features in polymers are on a scale larger than 2pm.

6.6.2 Measurement of the Thickness of an Isotropic Film Since in this example the film is isotropic, we are concerned with a single refractive index nf. We assume first that this is known. A small sample of film is selected and mounted between slide and coverslip as

214 D. A. Hemsley

FIG. 6.13. Method of mounting a film sample for thickness measurement. R and M show the intended paths of the

reference and measuring beams.

Fluid

Slide

R M

Coverslip

Film

shown in Fig. 6.13. The immersion fluid is chosen to bring the reference-specimen OPD within the measurement range of the compensator. Clearly this fluid should not swell, dissolve or otherwise attack the polymer film. Very thin film can be measured 'dry'. 1ft is the film thickness, the OPD is given by

OPD = tn L - tnr = t(nL - nr)

or

OPD (6.3) t =

where nL is the refractive index of the immersion fluid. If nr is not known, two measurements are obtained with different

immersion fluids of refractive indices n L1 and nL2' Then we have

t(nL1 - nr)

t(n~ - nc) (6.4)

Division of the two equations. allows nr to be calculated, and substitution into either equation gives the thickness t.

6.6.3 Measurement of Refractive Index of Spherical or Cylindrical Specimens

The OPD will vary across the specimen in accordance with the thickness variation. Figure 6.14 shows spheres of acrylic polymer, mostly displaying concentric circular interference fringes the spacing of which can be measured to prove that they are perfect spheres. Polymer particles that do not show a concentric fringe system are ignored in carrying out measurements of refractive index by this simple method.

If measurements of OPD are made at the centre of spheres, their diameters, measured with a micrometer eyepiece, can be taken as the


~ A o

FIG. 6.14. Spheres of acrylic polymer showing concentric ring fringe systems (X225). There are at least two species of particle represented which differ in their refractive indices. Type A shows more interference fringes for a given diameter

than type B.

thickness at the points of measurement, and the refractive index ns of an individual sphere is given by

OPD D

(6.5)

More generally, a plot of OPD versus D for several spheres has a slope giving the mean value of ns.

Similar measurements can be made on isotropic cylindrical fibres (e.g. in glass fibre composites) using a measured fibre diameter as the thickness in the calculation.

In the case of transmitted light fringe field interference systems a simple method of obtaining the OPD produced by a fibre is to measure the shift or 'deflection' of fringes covering the fibre at right angles (Fig. 6.15). If d is the deflection and s is the field fringe spacing, the OPD is given by dAis where A is the wavelength used. Since the OPD can be

FIG

. 6.1

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controlled by varying the immersion fluid, d can be kept within the field of view of the microscope.

6.6.4 Measurement of Refractive Indices of Anisotropic Thin Films and Fibres

Commercially produced polymer films generally show pronounced anisotropy of their physical properties arising from the molecular orientation introduced by the manufacturing process. Measurement of the optical anisotropy gives information which correlates with the degree and type of orientation and the other physical properties of the film. As described in Chapter 3, the optical anisotropy, or 'birefringence', can be used to characterize films and monitor properties. The birefringence, as measured in the plane of the polymer film, is the difference between two principal refractive indices (n, and n2) which can in principle be individually measured by interferometry. In practice the measurement of the individual indices rather than birefringence gives considerably more information about molecular organization.

To distinguish the principal indices it is necessary to use plane polarized light during measurement. Some types of microscope interferometer, e.g. the Zeiss (Oberkochen) Jamin Lebedeff system, utilize polarizing components in their construction and the waves passing through the reference area and the specimen are already plane polarized. In these instruments care is needed to rotate the specimen so that the required index is being measured. This clearly means that the user must be fully conversant with the characteristics of the interferometers. Other types of instrument, e.g. the Zeiss Jena Interphako system, normally operate with unpolarized light and a polarizer with a known vibration direction must then be added below the condenser unit and correctly orientated.

Suppose the thickness (t) of the plastics film is known and that specimen preparation is carried out as described above. For the first position of the film, i.e. when the direction of vibration of the light in the specimen part of the field of view is along the first principal axis, we have

(6.6)

On rotating the specimen through 90° we now have vibration of the light along the second principal axis and

(6.7)

218 D. A. Hemsley

Since t is known, each equation can be solved individually to give n I and n2' The birefringence, !!n, is given by (nl - n2)' A normal polarized light measurement may be used to check !!n directly.

If t is unknown, the double immersion method is used as before, but now with measurements for nl and n2 being taken for each fluid. Four equations result which can readily be solved for the three unknown quantities t, nl and n2' Indeed, one equation is redundant and may be used as a check.

The above procedure for films can be utilized also for fibres, and interference microscopy has long been an established method for fibre birefringence measurement8 and for the measurement of refractive index gradients.9

6.6.5 Measurement of Refractive Index of Phases in Sections The films and fibres discussed above require very little specimen preparation. However, if interferometry is to be carried out on bulk polymers such as mouldings or extrudates, these need to be thin sectioned prior to examination. Sections for interference work need to be of especially high quality with a minimum of knife marking or distortion.

Cross-sections of laminates or coextrusions reveal the constituent layers unless their refractive indices are extremely close. The actual OPD difference between the layers will depend on section thickness as well as refractive index difference, but third decimal place differences in index will usually be visible and measurable, with extension to the fourth place in favourable circumstances. Figure 6.16 shows a 20/lm thick section of a laminate of high density polyethylene (n~ = 1'530) and Nylon (n ~ = 1·535) with a thin polyurethane layer acting as an adhesive.

In calculating refractive indices from such cross-sections it is insufficient to assume that the sections are of the thickness indicated by the microtome setting. Two courses of action are available: either the double immersion method is used to determine both section thickness and the refractive indices or, alternatively, if one of the layers can be identified by other means (e.g. infrared ATR spectroscopy) then its refractive index is used to determine the section thickness which is then in turn used in calculations for other phases. This procedure is of particular use when a thick multi-ply laminate has its surface layers identified by other means but the identity of the inner layers is unknown.

FIG

. 6.

16.

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220 D. A. Hemsley

Multiphase polymers, such as plastics modified by the inclusion of rubber particles to improve mechanical properties, lend themselves well to microinterferometry. In practice, to be effective the rubber particle size is frequently well within the range visible with the light microscope. Measurements of particle refractive index may be used in identification of the rubber, and any boundary mixing effects can be studied.

Special interpretational problems arise when the rubber phase size is very small (say less than 5.um). Now, rubber particles will, in a thick section, not extend through the entire section thickness. Any rubber index measurement based on this assumption will be in error. However, the particles are often spherical, or can be persuaded to become so by mild thermal treatment of the section. Measurements can then be made along the lines described in connection with the polymer spheres. Alternatively, the section thickness could be reduced further, but this could result in small OPOs and a consequent loss of accuracy in measurement.

It is often unnecessary to carry out compensator measurements to check refractive indices. If a particular phase composition of known refractive index is suspected, then mounting the section in a liquid of this refractive index should give a zero OPO between the liquid and the phase under examination. This will occur regardless of section thickness, provided that the phase occupies the full thickness.

A more specific example of the use of transmitted light quantitative interference microscopy is its use in the study of systems in which polystyrene (Ps) was added to styrene-butadiene-styrene terpolymer (SBS) thermoplastic rubber. Earlier reports lO that there was limited solubility of the added Ps in the polystyrene domains of the SBS were checked using refractive index measurements on the SBS phase of the composite. The normal domain structure of the rubber is on a scale much too small to be resolved by the light microscope. As a result, the measured refractive index was simply a function of SIB ratio in the terpolymer. In fact the addition ofPs failed to produce any change in the SBS index,ll indicating that the molecular mass of the added Ps (substantially higher in the latter experiment) is significant in determining solubility.

6.6.6 Interference Microscopy of Liquids The refractive index of small amounts ofliquid can be determined using special microrefractometry slides. The amount of fluid necessary for


measurement amounts to only a few cubic-micrometres and the method has been used, for example, to identify fluid exuded by plasticized PVc. In this example the PVC contained a mixture of primary and secondary plasticizers and it was seen that, over a period of several weeks, traces of fluid appeared on the surface mouldings. Using microinterferometry it was possible to determine the refractive index of traces of fluid scraped from moulding surfaces. A knowledge of the refractive index assisted in identifying which of the plasticizers was moving to the surface of the product and whether its composition was constant over the period of exudation.

A microrefractometer slide consists of a high optical quality glass microscope slide with parallel flat surfaces and known refractive index (n g ). In the upper surface of the slide is a small cavity of regular and uniform dimensions. Often this cavity has the form of the cap of a sphere or a V-shaped groove. The depth of the cavity (D) is initially measured using normal microinterferometric methods. The liquid under investigation is then used to fill the cavity and a second measurement determines the new OPD between the cavity and a reference area. The refractive index of the liquid is given by

(6.8)

The ambiguity in sign is resolved by considering whether ng is greater or less than nL' This is in practice easily determined either by carrying out a Becke line test at the glass cavity walllIiquid boundary or by noting the behaviour of the interference fringes during compensator adjustment.

Measurements of the angle of contact between liquids and smooth polymer surfaces can provide useful data on the degree of wetting of the surfaces by liquids such as solvents. This might be of importance in formulating adhesive or coating systems. Alternatively such measurements may indicate the effectiveness, or otherwise, of chemical or electrical discharge treatments given to surfaces to enhance such surface properties as printing ink acceptance.

Although, in principle, the liquid/solid interface can be viewed using interference microscopy regardless of the angle of contact, in practice the technique is best limited to cases where the angle is small. A large contact angle gives a rapidly changing OPD over a short distance and an interpretable image can be difficult to obtain. However, small

222 D. A. Hemsley

contact angles allow accurate measurement of liquid layer thickness at the perimeter of a droplet, and measurements taken on a series of points close to the perimeter can be extrapolated to give a contact angle. 12

6.7 TRANSMIITED LIGHT SYSTEMS

As indicated by Francon5 and others,13 many transmitted light interference systems have been devised for the light microscope, but comparatively few of these reached the stage of commercial development and availability. It is to be regretted that the present trend is for fewer and fewer systems to be offered by the instrument manufacturers. The main reason appears to be lack of demand for what is a specialized and unavoidably expensive technique. Quantitative interference methods found favour with biologists in the 1950s and 1960s but their use has noticeably declined. This is not the case in other fields such as polymer microscopy. Here, as the above examples illustrate, there are many current applications for the technique and considerable scope for further utilization as interest, especially in polymer blends and composite structures, expands. Unfortunately the demand for instruments from this small (compared with biology) field of science and technology is unlikely to support a large interference microscope market or a wide variety of instrument types.

Below are described a number of systems currently available and, in briefest outline, others which may be found in established laboratories. The systems differ primarily in the method of beam splitting and recombination, the amount of 'shearing' (i.e. beam separation in the object or primary image planes) and the method of compensation for quantitative determination ofOPD in the field of view. Several systems, both past and present, have utilized polarizing optics to obtain beam separation and recombination, but the first system considered here does not, so for anisotropic polymer specimens it will usually be necessary, as we have seen, to incorporate a polarizer. A rotating stage on the microscope will also be an advantage when examining anisotropic specimens.

6.7.1 The Mach-Zehnder Interphako System This is the most versatile transmitted light quantitative interference system currently available and has a number of useful features not

To image plane


M - Z Interferometer

From lamp

223

FIG. 6.17. Basics of the Mach-Zehnder Interphako system: S is the specimen; o is the objective lens; RI and R2 are relay lenses; PI and P2 are the beam

splitting and recombination prisms respectively.

found in other instruments. In practice the system is capable of operation in a variety of different modes, including a differential mode used mainly for contrast enhancement. The system can also be used for reflected and transmitted interference work, but only the principles of the latter are considered here and they are further restricted to quantitative operation. We will also omit description of the interference phase contrast mode offered by the system since, although it can be employed as a quantitative method, it is restricted to small isolated particles showing small optical path differences from their surroundings. This type of object is quite uncommon in polymer work

A schematic outline of the system is shown in Fig. 6.17. The essential feature is that beam shearing and recombination both take place above the objective lens in a Mach-Zehnder interferometer. The physical design is greatly aided by the fact that the system employs 'infinity

224 D. A. Hemsley

corrected' objective lenses allowing the position of the primary image and other conjugate planes to be chosen by the designer. In use the system allows the lateral and variable shearing of the image by the production of two sets of beams each carrying all the specimen information. The phase relationship between these two sets is variable by means of a compensator (C).

A common mode of operation is to interfere the two beams so that, in the absence of a specimen, the 'background' interference colour is uniform across the field of view; this is known as 'homogeneous field' setting. Phase relationships across the field of view can also be varied to give a 'fringe field' image.

On recombination the two sets of beams interfere and in general one views a 'doubled' image showing lateral separation sufficient to superimpose the feature of interest and the reference background. This image doubling can be confusing if there is a concentration of features in the field (Fig. 6.18). Forthis reason the technique works best in 'dilute' fields with low feature concentration, or on isolated objects such as powder particles or fibres. In practice the amount of image shear is kept to a minimum consistent with full feature separation because large amounts of shear make greater demands on the coherence of the two waves and image contrast suffers.

Illumination of the specimen is by a normal condenser unit except that a slit aperture (A) is placed at its front focal plane - a position normally occupied by the aperture iris. The slit position in this plane (and its width) is variable and is set in relation to an interference fringe system observable in the back focal plane of the objective lens. The slit width controls coherence and has a marked effect upon contrast in the image of the specimen, so the system needs to be set up with care to obtain optimum results. Restricting the illuminating aperture in this way has consequences in terms of the resolving power of the microscope which is reduced in a direction perpendicular to the axis of the slit.

One of the primary advantages of this system is that (with the exception of the slit in the condenser unit) all the interference optics are contained in a single housing which can be removed and replaced readily to restore the instrument to normal working or to introduce alternative systems such as normal phase contrast or dark field.

Measurements of OPD are carried out by adjustment of the built-in compensator in the path of one of the beams in the interferometer head. Pre-calibration of the compensator is carried out using monochromatic light and fringe counting as the compensator is adjusted.

FIG

. 6.1

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226 D. A. Hemsley

FIG.6.l9. Basic construction of the PZO Pluta system.

6.7.2 The PZO Pluta System

To Eyepiece

--+--- Analyser

---+---Specimen

S----+-- - Polarizer

This system (Fig. 6.19) uses polarization optics to produce a shearing of the image above the objective lens. In practice, to obtain sufficiently large shear for quantitative work, this is carried out in two stages, first with a birefringent prism (PI) behind the objective lens (0) and then with a second (P2) in the body of the instrument. Again the system may be used under either 'homogeneous field' or 'fringe field' conditions and two laterally displaced images of the specimen are seen. Unlike the system described above, the amount of shear is fixed but the image appearance and interpretation are similar.

A built-in compensator system allows compensation of the two images for quantitative work. As above, the basic procedure in homogeneous field operation is to compensate first one image then the other; the difference in readings can then readily be converted into an OPD. In fringe field operation, fringe displacement may be measured to give an OPD but alternatively the fringes can be moved across the field by adjustment of the compensator to transfer a chosen fringe in the


background on to the specimen area of interest. Again a reading difference gives the OPD.

In either method it is again necessary to have a slit aperture (S) at the front focal plane of the condenser unit. As described above, the position of the slit and its width are set by simultaneous reference to an interference pattern in the back focal plane of the objective lens. The limitation on illuminating aperture imposed by the narrow slit widths necessary again degrades resolution in the image - at least in the shear direction. Nevertheless this is a useful system provided that the features for measurement are dilute in the field of view, present OPDs greater than aboutA/20 and are sufficiently large for the resolution shortcomings not to be prohibitive.

Since the system uses a polarizing system of beam control, this must be taken into account when analysing and carrying out measurements on anisotropic specimens. Close attention to the manufacturer's handbook is needed if interpretational errors are to be avoided with such specimens.

6.7.3 The Jamin-LebedetT System Based on a polarizing microscope, this system (Fig. 6.20) uses matched pairs of condenser and objective lens units. In the former, plane polarized light passing through the instrument is split with two orthogonally plane polarized beams (an '0' and an 'e' wave) by the use of a composite birefringent prism (PI). One of these beams eventually passes through a reference region (R) and the other through the specimen (S). The lateral separation between the beams is fixed in both size and direction for a given condenser/objective pair. For a X 10 objective/condenser this separation is typically 175 }.lm.

After passing through PI the beams pass through a half-wavelength (A/2) retardation plate (H). It can be shown that such a plate, with its axis at 45° to the vibration direction oflight passing into it, rotates the plane of polarization of polarized light passing through it by 90°. Since the beams from PI were originally orthogonally polarized, the effect of the A/2 plate is to interchange the vibration directions. On encountering a second prism (P2) identical to P], the original '0' wave behaves as an 'e' wave and vice versa. Recombination of the 'reference' and 'specimen' beams therefore takes place. As with a polarizing microscope, interference between the two plane polarized beams occurs when they encounter the upper polarizer or 'analyzer' which is normally in the 'crossed' orientation position with respect to the polarizer. Optical path

228 D. A. Hemsley

FIG. 6.20. Principle of the lamin-Lebedeff microinterferometer.

l /

R

H

l V

Ana Iyser

/ I

S

I

Pol arizer

differences between the beams passing through the reference and specimen areas therefore show up as differences in light intensity (monochromatic light) or in polarization colours (white light).

The system is relatively easy to use and has found many applications in polymer work. One of the advantages is that compensators used for polarized light measurements can be used also for OPD measurements in the transmitted light interference mode.

Since the A12 plate will perform as described only for a specific wavelength (A), strictly the system can only be used with monochromatic light. However, white light images are suitable for qualitative work and for determining fringe order. Normally A = 546 nm (green light), so for the blue and red ends of the visible spectrum the performance oftheM2 plate is imperfect and some components oflight pass on to P2 with the original vibration directions determined by PI as shown in Fig. 6.3. It is therefore possible to determine the 'reference' region of the field of view since this has a faint magenta (red + blue) colour. Under the usual conditions of Kohler illumination, partial closing of the field iris until


the magenta area andthe specimen area of the field of view just separate ensures no overlap of 'reference' and 'specimen' imaging beams. In practical use this adjustment proves to be a significant advantage of this system since one can readily assess the content of the reference area which must of course be kept clear of specimen during measurements.

6.7.4 Other Types of Transmitted Light Interference Microscope Two other instrument types deserve mention although they are no longer commercially available. Their inclusion here is justified because they have special merits, and applications exist for which they are especially suitable.

The first of these is the 'double focus' instrument based on a design by Smith and described by Francon.5 Instead of the incident beam being split laterally, it is arranged (by using polarizing optics) that two images of the specimen are formed which are separated vertically. The light coming from any point of the in-focus image is made to interfere with light forming the out-of-focus image. The phase of the light from any specimen feature is thereby compared with the phase of the light from the surrounding area of the field of view. This system, although capable of being used in a quantitative mode, is primarily used for qualitative microscopy. It has the advantage that the objectives can work at their full NA but competes with differential interference systems and phase contrast. The biggest advantage over these techniques is that a feature having a constant optical thickness will appear uniformly bright or coloured. Compared with the other interference systems described above, the major advantage is that only a single image is in focus. Putting these two advantages together, one sees that the image presented is easily 'interpreted' by automatic image analysis equipment. With increasing interest in such equipment it is unfortunate that the most suitable interference microscope for such work is no longer available.

The second instrument of interest, and one briefly described by Roche and Davis,9 was also based on the Mach-Zehnder interferometer. Unlike the instrument described in Section 6.7.1, beam splitting took place, by the use of prisms, at a position below the condenser unit, and recombination was allowed just below the eyepiece of the microscope. In effect the system was a double microscope with carefully matched optical components. The primary advantage of this equipment was the wide beam separation - no less than 60 mm! This allowed large polymer specimens to be examined at full field and objective apertures.

230 D. A. Hemsley

6.8 COMPENSATORS

As mentioned earlier, it is usually possible subjectively to assess the magnitude of OPDs between about ).J2 and 3}" by observing the interference or polarization colour shown by a specimen. More objective and precise measurements are carried out using compensators. Compensators fall into two basic types according to whether or not the interference microscope uses polarization optics.

In cases where this is not so, e.g. the Mach-Zehnder designs, the problem is simple because the beams follow different well separated paths in the region where a compensator can be placed. The relative optical path lengths can be changed simply by arranging for one of the beams to traverse a thickness of glass that can be varied by tilting or sliding a wedge. The 'reference' beam is made to go through a thickness of glass equivalent to the centre thickness of the compensator so as to allow the latter relatively to advance or retard the beam.

Systems based on polarized light optics use standard birefringent compensators such as would be used in birefringence measurement (discussed in Chapter 3). Sometimes the compensator is dedicated to the interference system, as in the Pluta instrument and the Smith 'double focus' system. Alternatively, standard slot-in compensators are used; these may be of the Ehringhaus, Berek or Elliptic type according to the magnitude of the OPD being measured, the latter being for small path differences. De Senarmont compensation may also be used for small path differences.

Birefringent compensators are positioned at a point in the optical system where two beams orthogonally polarized pass through the microscope, and the aim is to adjust their relative phase difference to zero. Just as in birefringence measurement discussed in Chapter 3, dispersion problems arising from differences between the refractive index versus wavelength curves for the specimen and the compensator can occur. The possible solutions to the dispersion problem are also the same and are not discussed further here.

Regardless of compensator type, precision of measurement can be enhanced by the use of devices to allow more accurate setting of the compensator. Without such devices one is normally setting, by eye, a part of the field of view to minimum intensity or to a specific colour.

The accuracy achieved this way may be 1/200 of a wavelength using monochromatic light; the limitation is the ability to judge the colour or the spot of maximum extinction (particularly for small features) accurately by eye. This can be overcome by optical means, by introducing


a 'half-shade plate'. This plate is a phase step of usually 1/4 of a wavelength and divides simultaneously the feature and background into two parts. It is placed in an intermediate image plane and is therefore visible with the image of the specimen. By superimposing this constant phase difference on both - half on the feature and half on the background - one can set the variable phase of the light going through the compensator to the very critical point of equal phase difference (and light intensity) between the divided parts of the feature or the background. Readings by this route can result in accuracies of up to 1/500 of a wavelength, but even that can be improved by electronic means.

Ultimately one can have confidence in measurements of refractive index of2 X 10-4 in a 10 pm thick section or a thickness measurement of 2 pm for a O' 5 refractive index difference.

REFERENCES

1. Hartley, G., Hartley's Microscopy. Senecio Publishing Co., Oxford, 1978, p.152.

2. Hecht, E. & Zajac, A., Optics. Addision-Wesley, London, 1974. 3. Francon, M., Optical Image Formation and Processing, Academic Press,

London, 1979, p. 66. 4. Born, M. & Wolf, E., Principles o/Optics, 2nd edn. Pergamon Press, Oxford,

1964. 5. Francon, M., Progress in Microscopy. Pergamon Press, Oxford, 1961,

pp.94-128. 6. Tolansky, S., Multiple Beam Interferometry 0/ Surfaces and Films. Oxford

University Press, 1948. 7. Hemsley, D. A., The Light Microscopy o/Synthetic Polymers, RMS Handbook

7. Oxford University Press, 1985. 8. Hamza, A. A. Optical birefringence phenomena in fibres. Textile Research J.,

53(4) (1984) 205-9. 9. Roche, E. J. & Davis, H. A., Measurement of radial birefringence in fibres.

Fiber Producer, 51 (1984) 51-5. 10. Skoulios, A., Helffer, P., Gallot, Y. & Saleb, J., Solubilization and chain

conformation in a block copolymer system. Makromol. Chem., 148 (1971) 305-9.

11. Nandra, D. S., Hemsley, D. A. & Birley, A. W., Reinforcing resins in SBS copolymer elastomers: properties and microstructure of some injection mouldings. Plastics and Rubber: Materials and Applications, 4 (1976) 38-43.

12. Longman, G. W. & Palmer, R. P., Two microscopical methods for determining the contact angles of small drops. J. Colloid and Interface Sci., 24(2) (1967) 185-8.

13. Krug, W., Rienitz, J. & Schulz, G., Contributions to Interference Microscopy. Hilger and Watts, York, 1964.

7

Ultraviolet and Fluorescence Microscopy

P. CALVERT and N. C. BILLINGHAM

School a/Chemistry and Molecular Science, University a/Sussex, Brighton, UK

7.1 INTRODUCTION

The optical system of any microscope can be divided into two main groups or components, the illuminating optics and the observing optics. The two systems are designed so that, in the absence of a sample, the field of view imaged by the observing optics appears uniformly illuminated (or uniformly dark). When a sample is introduced into the microscope, image contrast arises because different regions of the sample affect the illumination to differing extents. The image may result from many effects, including diffraction, refraction, reflection, scattering, interference, polarization, fluorescence and absorption, and microscopes have been designed to take advantage of most of these phenomena. In this chapter we shall be concerned with optical microscopy in which the illuminating light is in the blue/ultraviolet range, from around 230 nm to 400 nm. In this range the main intentional sources of image contrast are uv absorption and fluorescence emission, although other mechanisms, notably diffraction, may cause problems.

Fluorescence emission is normally observed in the visible, and the complex and expensive observing optics of the microscope can therefore be constructed of conventional optical glass. The fluorescence microscope has become a routine tool of great power in biological and medical science, but its use in studying synthetic polymers is rare. In contrast, UV absorption microscopy requires much more sophisticated equipment since the observing optics must be able to transmit UV

radiation, and some method is required for shifting the image into the visible for focussing and observation.

233

234 P. Calvert and N C. Billingham

The uv microscope was originally developed by Kohld in order to take advantage of the increased resolving power theoretically associated with shorter wavelengths. The increased resolution actually yielded little new information but the microscope did show unexpected contrast effects in biological samples which were completely transparent in visible light. It was later shown that these effects are due to the strong absorption of UV radiation by nucleic acids, and this observation quickly led to extensive use ofuv microscopy to study the distribution of nucleic acids within cells. The development of the electron microscope has meant that there is little advantage in using UV light to obtain increased resolving power, as compared with the enormous increase allowed by electron microscopy. Rather, most uses have been to make qualitative or quantitative concentration observations on systems where one component is strongly UV absorbing.

Most commercially important synthetic polymers have no strong UV

absorption in the easily accessible range from 250 nm to 400 nm, nor do they contain fluorescent centres. Hence useful application of the uv microscope will depend on there being added uv absorbing molecules or attached side groups whose concentration varies within the polymer. Since the only systems that obviously fall into this category are polymers containing UV stabilizers or optical brighteners, this has until recently been the only application in polymer science. It has been our belief that the potential range of applications is very much wider than this, in that UV absorbers or fluorescers can be selectively bound to specific chemical entities in the polymer or will preferentially interact with, or dissolve in, parts of the structure. In this way these molecules can be used as stains and probes of the morphology of the polymer on the scale from 0·25 pm upwards, in a manner very similar to that in which the biologist uses stains to develop contrast in tissue specimens. Further, insofar as UV absorbers resemble other small molecules of interest, such as drugs and pesticides, they can be used to study the transport of such molecules in polymers. In principle, similar measurements could be made with coloured substances using a normal visible light microscope. However, in all forms oflight microscopy the depth of focus is limited, particularly as the magnification is increased. The result is that very thin samples are required for successful light microscopy, and only absorbing species with high extinction coefficients will yield acceptable contrast. The main advantages ofuv illumination are thus the greater range of absorbing compounds with a high extinction coefficient and the number of UV absorbing substances that are of interest in their own right. A further advantage of the uv

Ultraviolet and Fluorescence Microscopy 235

microscope is that it can be used as a fluorescence microscope, although the reverse is not true. Observation of fluorescing substances can offer greater sensitivity since the fluorescence is observed against a dark background, but the range of suitable compounds is more limited.

Work carried out up to 1981 on the development of UV and fluorescence microscopy for the study of synthetic polymers was the subject of an earlier review.2 In this chapter the aim is to summarize, update and extend that review. We concentrate on applications rather than on a detailed description of experimental methods, since these were described in full detail in the earlier review.

7.2 EQUIPMENT AND TECHNIQUES

7.2.1 Microscope Design In this section we describe the basic requirements of a microscope for uv and fluorescence work, and review some recent developments in instrumentation.

In essence UV illumination requires a suitable light source, whose wavelength can be selected to be appropriate for the absorption maximum or excitation wavelength of the species being observed, and an optical system that can transmit and focus the radiation. For fluorescence work the observing system must be capable of forming an image from the visible emission from the sample. In uv absorption work the observing system is required to transmit radiation in the uv and it is necessary to have some means of presenting the image in the visible since the wavelengths used are damaging to the eye.

Figure 7.1 shows, schematically, typical equipment for the uv and fluorescence microscopy of polymers. It is a normal Zeiss Universal microscope which has been adapted for UV illumination. All of the optical system is quartz, using Zeiss Ultrafluar objectives and condenser, which are achromatic over the range 220-700 nm. This is a most desirable feature since it allows focussing of the microscope in the visible with the image still being in focus in the uv. It also means that resolution is not lost if the illuminating light is polychromatic. Front surface mirrors must be used in all the beam switches. The light source in our particular instrument is a 150 W xenon arc lamp, with wavelength selection by interference and coloured glass filters. The main requirement of the illuminating light in UV work is that its wavelength distribution should lie entirely within the absorption envelope of the

236 P Calvert and N C. Billingham

WAVEFORM r--- PICTURE

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FIG. 7.1. Block diagram of the UV microscope.

sample, preferably with a sharp spectrum at the absorption maximum, so as to maximize image contrast. For fluorescence work the light should be confined to the excitation spectrum of the sample, and radiation in the emission range should be filtered out. These requirements are usually not too severe and can be met quite easily by using filters. There is little advantage in using a monochromator, and it is a positive disadvantage for fluorescence where the weakness of the


emission requires maximum illumination intensity. The illumination is operated as closely as possible under Kohler conditions and must be set up carefully to ensure uniform illumination of the field of view, especially for quantitative work.

For viewing the transmitted UV image it is most convenient to use a TV camera, fitted with a quartz fronted, uv sensitive, tube. The signal from the camera is viewed on a TV monitor and can also be analysed by a waveform monitor. This latter is a very useful instrument which selects any line on the TV picture and displays a trace of its white level on an oscilloscope. It thus acts as a microdensitometer and is used for monitoring the uniformity of illumination when setting up the microscope, and for making quantitative measurements of absorption or fluorescence. In addition, the image can be switched to a conventional 35 mm camera for permanent recording, and a separate camera allows recording of the waveform monitor screen. For fluorescence work we fit a barrier filter above the objective to remove excitation wavelengths. In addition, the microscope has a tungsten light source and a normal eyepiece, so that it can be used for conventional microscopy, and it is equipped with polarizing and analysing filters.

Such a system represents the minimum requirement for successful uv absorption microscopy. If fluorescence microscopy is the only facility needed, the requirements are simpler and the range of microscopes available is much greater. In transmitted fluorescence the simple Abbe condenser of a normal microscope is usually replaced by a dark field condenser whose smallest numerical aperture is chosen to be larger than that ofthe objective. In ideal conditions the exciting radiation does not then reach the objective, and only the fluorescence emission can contribute to the image. In practice, exciting radiation is diffracted by simple irregularities, and also enters the objective, but it can easily be filtered out above the objective. In a transmission fluorescence instrument the condenser is set up to illuminate the area of the sample viewed at the lowest magnification. As the magnification is increased the area viewed decreases, so the total energy available for forming the image is reduced. This factor has led to the development of an alternative system in which the illumination is by reflection. In a typical reflection fluorescence microscope the light source is mounted above the sample plane. The source light is incident upon a dichromatic mirror which reflects the short wavelength exciting light through the objective, which focusses it on to the specimen plane. Any light that is not absorbed passes harmlessly through the specimen and does not

238 P. CaJvert and N C. Billingham

enter the imaging system. The specimen fluorescence is captured and imaged by the objective. The fluorescence, being of longer wavelength, is transmitted by the mirror which reflects any diffracted components back to the source. A barrier filter is usually employed to increase image contrast. Because the objective is used as a condenser, only the area of sample being viewed is excited, giving improved sensitivity at higher magnifications. In addition, the absence of a separate condenser reduces alignment problems. In reflected fluorescence, illumination and imaging take place on the same side of the specimen so the maximum fluorescence is produced in the specimen layer being observed. Reflected fluorescence generally gives brighter images, especially from thick samples, and it is much more popular in routine fluorescence microscopes.

Although the basic microscope is perfectly satisfactory for most work with polymers, far more elaborate systems have been produced; a number of these have been reviewed by Freed.3 If the light source is fitted with a scanning monochromator and a photomultiplier tube is fitted to the observing system, the UV microscope becomes a microspectrophotometer and can be used to measure absorption spectra on small areas of a specimen and to make quantitative measurements of concentration. This approach has been used to look at stabilizing additives in polymers4 and has also been explored in studies of biological cells.5 The main problem is sensitivity,6 since the small depth of field of a microscope operating at high magnification limits the sample thickness to a few micrometres, so only species with very high extinction coefficients can be examined. In contrast, fluorescence is potentially up to four orders of magnitude more sensitive than absorption; fluorescence spectrometry is a very powerful analytical tool for fluorescent species. Fluorescence microspectrophotometry requires monochromators both above and below the specimen, together with a suitable photomultiplier system. Such systems have been discussed/ and Gurkin and Kalld describe a commercially available instrument capable of recording fluorescence excitation spectra from areas of a specimen as small as 1 pm in diameter and which can be used for quantitative measurements of fluorescence intensities.

The strong absorption of UV at 260 nm by nucleic acids allows UV microscopy to be used to study the distribution of DNA and RNA in whole cells. This approach has been extended to allow kinetic studies of living cells at different stages in their growth cycle and to study the response of cells to different treatments. Special microscope systems


have been developed to avoid the killing of cells by the high UV exposures normally needed. These applications have been very extensively reviewed by Freed3 and by Blout.9 Because of the high intensity ofuv required, the usual method is to synchronize scanning of the UV beam with the scanning of the TV camera used to observe the image.

Normally a conventional TV camera is used for viewing the image but some workers have used more sophisticated approaches. Image intensifiers provide increased sensitivity at extra cost and provide a visible image which can be photographed. For fluorescence microscopy an alternative approach has been to combine an image intensifier with a TV camera to produce a 'video intensification camera' (VIC) and several authors 10. II have commented on the value of such cameras in fluorescence microscopy. Willingham and Pastanl2 combined the VIC method with a time-lapse video recorder for examination of live cells, claiming the sensitivity obtained from the VIC was high enough to eliminate the need for scanning systems. More recently, Forman and Turriffl3 have commented on the advantages of this approach for more conventional fluorescence microscopy of fixed cells; in particular, the use of a TV image eliminates the need for dark adaptation while using the microscope.

Another sophisticated approach to live cells has been described by Inoue. 14 In his system the same sample is viewed simultaneously by two different intensified cameras. One camera records the fluorescence image and the other a polarized light or differential interference contrast image. These two images are then combined in real time and in different colours, and the composite image is video recorded. In this way the weak fluorescence image from a living cell can be related to any birefringence or optical path difference in the cell. It is claimed that this is a powerful method for the study ofliving cells without UV damage.

The most intense sources of monochromatic radiation currently available are lasers, and it is not surprizing that they are beginning to appear as light sources for microscopy. In our own work we found some advantages in using a helium-cadmium laser as the illuminating source for UV microscopy, but the high cost ofuv lasers limits their use to more sophisticated applications. In 1961 Ambrosel 5 described a microscope designed to observe the contact region between growing cells and the surface on which they are growing, by using illumination generated by radiation totally internally reflected at a water/glass interface. Under such conditions the internally reflected wave actually penetrates the


water (to a depth of about 20 nm for visible radiation) and can excite fluorescence in this contact region. The effect is extremely weak and contrast low. Recently Axelrod et al. 16 have adapted this technique by using an argon ion laser, operating at 514·5 nm, and claim to be able to observe fluorescence from cell membranes with much reduced contributions from internal structures.

Perhaps the most sophisticated fluorescence microscope yet developed is the time-resolved fluorescence system described by Docchio et al. 17 In this system a conventional incident fluorescence microscope is illuminated by a pulsed dye laser which is in its turn driven by a nitrogen laser. The combination illuminates the microscope with subnanosecond pulses of around 10 kW peak power, tunable from the near uv to the near infrared. The area of sample excited into fluorescence is of the order of o· 3 flm in diameter. Light emitted from the sample is collected by the objective, filtered to remove laser reflections, and detected by a fast photomultiplier tube. A microprocessor system allows analysis of the decay of fluorescence after the excitation pulse, and hence the determination of fluorescence decay times in the nanosecond range. This decay time is a function of the environment of the fluorescing centre and can be used to probe interactions of these centres with their surroundings. This method has been successfully applied to study the distribution of fluorescent drugs in cells, and the nature of their interactions with the cell membranes.

7.2.2 Sample Preparation In an ideal microscope the sample plane is uniformly illuminated by the sub-stage optical system. Contrast in the image then arises from any effect that non-uniformly reduces the light reaching the objective. For solid samples with rough surfaces, a particular problem is the contrast produced by diffraction effects at the upper and lower surfaces, and some care is required in sample preparation if artefacts due to diffraction are to be avoided. Sample preparation for uv microscopy is described in detail in Ref. 2; only the important points are summarized here.

For work with polymers, samples are generally in the form of microtomed slices of 5-10 flm thickness (see Chapter 1). We section samples with a base-sledge microtome, using glass knives and a cooled stage. The sections are mounted between a slide and a coverslip, both of which must be of quartz for work below 350 nm; these are readily available but expensive and regrettably fragile. As an alternative to slices, solvent cast


films may be prepared by dripping a dilute solution of the polymer on to a heated slide. If the sample permits, it is fused to the slide and coverslip by rapidly heating them on a hot-plate pressing. Ifthe sample cannot be fused to the slide, a mounting fluid must be used to reduce the diffraction contrast. The requirements for this fluid are that it should not swell or otherwise interact with the polymer, should not extract the additive, should not be uv absorbing, and its refractive index should match that of the polymer. Most polymers have refractive indices around 1· 5, and wholly satisfactory mounting fluids are rare. In particular, virtually all non-extractive oils with refractive indices above 1-48 are too strongly UV absorbing for use in the uv; generally glycerol is used.

The existence of diffraction contrast may be assessed by comparing visible and UV images of the same sample, particularly adjacent to a sample boundary. An obvious darkening of the sample with respect to the surrounding liquid in the uv, with no similar darkening in the visible, is good evidence for absorption contrast. In fluorescence microscopy the absence of a close refractive index match leads to a bright line, seen around the edge of the sample and on internal surfaces. This arises from light radiated within the plane of the sample and reflected at low angle from the top and bottom surfaces such that it finally emerges at the boundary.

7.2.3 Quantitative Measurements For many applications of the uv microscope it is necessary to make quantitative measurements of the concentration of UV absorber as a function of position within the field of view. A convenient method for concentration measurement in UV microscopy is to apply the output of the TV camera to a wavefonn monitor which displays the intensity along any selected line of the TV image. The monitor cannot normally be applied in fluorescence work, as the intensities are too low to give a TV picture; for fluorescence work it is necessary to use photographic methods or image intensifiers. The waveform monitor requires calibration for quantitative use. The methods of calibration are essentially the same as those used in photographic work and described below.

An alternative method of making concentration measurements is to use scanning microdensitometry of photomicrographs. Microdensitometry of photographic images has been discussed in detail by many people but it is useful to summarize the basic arguments. In UV


absorption measurements the light intensity transmitted by the sample is governed by the Beer-Lambert law. The response of the photographic film to light is ideally to produce an optical density D such that the film transmittance is inversely proportional to the intensity oflight reaching the film (the reciprocity law). Hence the difference in optical density between a transparent part of the sample of thickness I and a UV

absorbing part is given by

Do - D oc eel (7.1)

so the optical density of the film is directly proportional to the absorbance of the sample. For UV absorbers the analysis is thus most conveniently done by scanning the negative with a double-beam recording microdensitometer. This instrument yields absorbance values which are directly proportional to the optical density of the film. Scott et al. 18 and Freed3 have discussed the necessary corrections in detail. Care must be taken in exposing the film to avoid reciprocity failure and to process all films under identical conditions. Ideally, calibration and sample exposures should all be on a single roll of film. Calibrants are generally a series of samples with known UV absorber concentration.

In fluorescence, the emitted intensity depends upon the amount of exciting light absorbed and the quantum efficiency for fluorescence of the excited state, leading to an expression of the form

I = loB [1 - exp( -eel)] (7.2)

This expression can easily be analysed only where the absorbance is small, when

I/Io oc eel (7.3)

Thus measurements of film transmittance should be proportional to concentration. For fluorescence work a single-beam microdensitometer may be used which gives transmittance readings directly.

7.3 APPLICATIONS TO NON-POLYMER MATERIALS

As described in the introduction, fluorescence microscopy has been developed very extensively in biological and medical science, aided by the development of a large range of staining reagents capable of selective binding to structures of interest within cells. There are many


variations of this technique, designed to allow studies of living cells without damage, and some of these have been described above. In contrast, uv microscopy has received rather less attention, mainly because of the greater expense of the optics required and the lower inherent sensitivity. Both uv and fluorescence microscopy have found applications outside straightforward cell biology, and it is these applications that are reviewed in this section, since it is useful to be aware of uses of this technique on other materials, as a guide both to what is possible and to the experimental methods that can be used.

Hass and Plath19 used a uv microscope spectrophotometer to study single crystals of zeolites having varying contents ofNi{II) and were able to determine the symmetries of nickel complexes within the zeolite structures. They claim that the spectra obtained by them are very much better than any from previous attempts using powder reflection methods.

In addition to simple absorption measurements microscopic circular dichroism has been developed for biological work. 20 Dichroism measurements with polarized uv have been used to look at anisotropic arrangements of nucleic acids in chromosomes, viruses and sperm.9 The anisotropy of muscle fibres has been studied by observing the polarized uv fluorescence from tryptophan.21

A major area for the use of UV microscopy is in the observation of lignin in wood as described by Goring and his co-workers. They have measured spatial distributions of lignin in various woods,18.22-24 the effects of chemical treatments in lignin extraction,25 and separate distributions of syringyl and guaiacyl residues.26 Fluorescence microscopy has been used to observe the lignin precursor, ferulic acid, in cell walls of grasses and cereals.27

An alternative method for estimation oflignin in wood is bromination, followed by energy dispersive X-ray analysis in the electron microscope.28 There have been reported discrepancies in the results from these two techniques, and Saka et al. 29 have looked at the reasons. They find that the uv extinction coefficient for lignin is generally independent of the time at which the lignin was formed in the cells; the reactivity towards bromination is significantly variable with the location of the lignin, so UV microscopy is regarded as the more reliable technique. Fukazawa and Imagawa30 have also discussed the application ofuv microscopy to quantitative determination of lignin.

A Russian review has described applications of UV microscopy in mineralogy.31 One particular area of application is in coal and oil

244 P. Calven and N C. Billingham

petrology. Reflected light fluorescence microscopy is used, primarily with 365 nm excitation, to measure the organic content of coal and peat.32 Reflectivity in the visible and ultraviolet can also be related to the carbon content of coals.33 Transmission uv microscopy of coal has been described once, but it requires ultra-thin sections.34 A bibliography of coal and oil petrology has been prepared by Zeiss.35 Teichmuller and Durand36 have described the use of fluorescence microscopy to rank coal structures, whilst Shibaoka and Russell37 have shown that fluorescence microscopy can be of use in the study of residues from coal hydrogenation. Crellini8 has recently reviewed the use of fluorescence microscopy in coal petrology.

7.4 NON-MICROSCOPIC APPLICATIONS OF FLUORESCENCE FROM POLYMERS

Fluorescence is a phenomenon of more general scientific interest than absorption, for several reasons. The fluorescence itself can be observed with much greater sensitivity than absorption, for appropriate species. The fluorescence emission from an excited molecule decays with a relaxation time that is sensitive to the local environment of the fluorescing centre, and the emission may be polarized in a way that is revealing of orientation in the sample.

In our own work we have used UV absorbers and fluorescers in polymers to give both images and concentration information. Fluorescent markers have also be used by others to study mobility, orientation and phase behaviour in polymers, and there is no reason why the same methods could not be applied in conjunction with microscopy if the system warranted it.

There are recent reviews of fluorescence methods for polymers by Monnerie/9 Morawetz40 and Nishijima.41 In solid polymers and polymer melts it is possible to measure molecular mobility by using fluorescent probes which may be attached to the polymer or simply dissolved in it. Data can be obtained either by exciting the fluorescence with polarized light and measuring the extent of depolarization of the emission as a function of temperature, or by exciting the probe molecule with a short pulse of light and monitoring the time dependence of the depolarized emission. Different mobility ranges can be studied by using probe molecules with different fluorescence lifetimes. If the probe molecule is an elongated rod-like species, it may take on the same


orientation as the polymer molecules surrounding it, and can be used to study orientation in the polymer.42. 43 The fluorescence emission is measured as a function of the sample orientation, using crossed polars, and can be used to compute the second and fourth moments of the orientation distribution.

Recently fluorescence has been used to study phase separation in blends of polystyrene and poly(vinyl methyl ether).44 Anthracene groups attached to the polystyrene chain fluoresce efficiently, but their fluorescence is quenched by the ether functions in the other polymer. Phase separation reduces these quenching interactions and is accompanied by a sharp increase in emission intensity. The quenching of the fluorescence of a molecule in a polymer by other species has also been used to monitor the diffusion of small molecules, such as oxygen, into polymers. A recent development of this method allows monitoring of polymer self-diffusion.45 In this method, poly(propylene oxide) was labelled with one dye molecule per chain and mixed at around 0·1 % concentration in unlabelled polymer. The dye fluorescence was locally and irreversibly photobleached by a laser pulse, and a second laser was used to monitor the diffusion of fluorescent species into the photobleached region.

Lamarre and Sung46 have used a related technique to study molecular mobility and physical ageing in amorphous polymers. In their method, azobenzenes are incorporated into various parts of the polymer chain. Both the trans-cis photoisomerization and the cis-trans thermal isomerization can be monitored spectroscopically. Since the isomerization involves a substantial change in the conformation of the probe, its rate depends strongly on the mobility of the polymer in the region of the probe molecule.

7.5 APPLICATIONS OF UV MICROSCOPY TO SYNTHETIC POLYMERS

Synthetic polymers do not usually have absorptions in the near uv, nor do they contain strongly fluorescent centres, so pure polymers do not exhibit any contrast when viewed in the UV or fluorescence microscope. In order to develop contrast it is necessary to have such centres distributed in the polymer in such a way that they reveal features of interest when viewed in the microscope. The observable centres may be present for a number of reasons: (1) they may be adventitious impurity

246 P Calven and N C. Billingham

groups present in the polymer or induced by the degradation or other reactions of the polymer during processing or service; (2) they may be deliberately added in the form of soluble but non-reacting additives whose solubility in the polymer is influenced by density variations and crystallinity so that they can be used to reveal density fluctuations or morphological features in the polymer or to monitor migration of the additives in the polymer; (3) they may be added deliberately as reagents capable of reacting chemically with features of interest, such as sites produced by degradation, so as to reveal the distribution of otherwise invisible sites.

7.5.1 Fluorescence from PVC When PVC is processed or exposed to outdoor weathering, the major mechanism of degradation is dehydrochlorination by elimination of HCl. This process is autocatalytic and produces conjugated sequences of double bonds in the polymer backbone, with consequent discolouration of the polymer. It is known that these sequences are able to fluoresce if exposed to near uv radiation. Hemsley et a1. 47 used fluorescence microscopy to monitor the degradation reactions during handling and processing of PVC powders. They found that the fluorescence produced in powder grains by mild thermal treatment can be used to monitor the fate of these grains during processing of the polymer by dry blending or extrusion, and they were able to monitor the dispersion of stabilizer by its ability to remove the fluorescence.

7.5.2 Orientation Studies Pinaud et a1. 42 have described the use of fluorescence polarization microscopy to study the amorphous orientation in polypropylene. They used a fluorescence microscope modified to allow polarization of the incident light and measurement of the polarization of the fluorescence emission. All-trans-l ,8-diphenyloctatetraene was used as the fluorescent probe molecule. They found that, when the depolarization effects due to scattering are properly corrected, fluorescence microscopy provides valuable information on the amorphous orientation, as a function of time and position during drawing. The work on polypropylene suggests that amorphous chains are not free to orientate or disorientate independently of the crystallites when the polymer is deformed. Instead their orientation is strongly correlated with that ofthe crystal phase and largely determined by the crystalline orientation and morphology.


7.5.3 Rejection of Impurities during Polymer Crystallization Starting from an interest in the action of light stabilizers and antioxidants in crystalline polymers, we have made extensive studies of the behaviour of impurities during polymer crystallization. With the uv microscope it is possible to measure small «1%) concentrations ofuv absorbers or fluorescers in a polymer. As the polymer is crystallized impurities are generally able to dissolve only in the amorphous phase, so they will become redistributed by the growing spherulites and the consequent concentration variations can be monitored by the uv microscope.

Figure 7.2 shows a typical uv micrograph of a fully crystallized sample of polypropylene containing a uv absorber and clearly demonstrating the rejection process. This rejection is not especially surprising since it is well established that even molecules as small as

FIG. 7.2. Polypropylene containing 0·5% Uvitex OB, crystallized at 125°C and viewed in transmitted UV light. Bar = lOO.um (X 160).


oxygen are unable to enter the crystal phase of polypropylene.48 Further, molecular models show that typical UV absorbers have dimensions comparable to those of the unit cell of polypropylene and are therefore unable to enter the crystal lattice without disrupting it. The theory of impurity partitioning in solidifying systems was first evolved for the solidification of metals49 and later became important in zone refining of semiconductors.5o We were able to show' that small molecules in polypropylene partition according to a simple model in which the additive is totally rejected from the crystal phase but remains dissolved in the amorphous material both within and outside the spherulites. In these systems the spherulite can be modelled as a uniformly growing sphere with a partition coefficient for the additive equal to the fraction of amorphous material in the spherulite (assuming that amorphous polymer within and outside the spherulite has the same dissolving power for the additive). The additive is assumed to move through the amorphous polymer only by diffusion, and convection is negligible in polymer melts. Observations were made of additive concentration gradients around growing spherulites in polypropylene containing 0·1-1·0% of a number of benzophenones, phenolic antioxidants and Uvitex OB, a fluorescent optical brightener. Results from observations both during spherulite growth in a hot stage and in samples quenched during crystallization and subsequently sectioned were fitted to computed distributions for the concentration of a rejected additive around two- or three-dimensional spherulites growing into a melt (Fig. 7.3). The measured crystallinity of the spherulite (from scanning calorimetry) can be compared with that derived from the concentration step at the spherulite interface (Table 7.1), and the diffusion coefficient of the additive in the liquid polymer is deduced from the shape of the concentration gradient in the liquid ahead of the growing interface (Table 7.2).

In more general terms, the behaviour of an additive in a polymer will depend upon its solubility in the crystal and amorphous phases, its diffusion coefficient compared with the growth rate of the spherulite, the sizes of the individual propagating crystals, and the quantity of additive present. Most of the additives studied were deliberately chosen to be soluble in the amorphous polymer at the concentrations used. In one case, Nonox CI, a phenolic antioxidant, precipitation of the additive initiated a high concentration zone around the spherulite, leading to a ring of precipitate. Preliminary work on totally insoluble particles has not shown any signs of rejection, although this has been seen in crystallization of low molecular weight compounds.52 In


~135 u 135

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(a) (b)

FIG. 7.3. Distributions of Uvitex OB around polypropylene spherulites at 125° C, 130° C and 135° C. (a) Intensity traces from transmitted UV images. The central, high intensity regions (low Uvitex concentration) are diameters of growing spherulites. (b) Computed relative concentration distributions for Uvitex OB in polypropylene calculated with a diffusion coefficient of8 11m2 S-I.

copolymers and partly degraded polymers, 'impurity' species may be partly incorporated into growing crystals. Rejection of low molecular weight polymer in polyethylene has been studied by extraction procedures.53

The characteristic distance scale for a rejected impurity is given by the ratio.(D/G) of the diffusion coefficient to the growth rate of the spherulite. For most of the compounds studied in polypropylene this


TABLE 7.1 Rejection of Additives and Crystallinity in Polypropylene

Crystallization Interfacial Crystallinity from Crystallinity from temperature partition coefficient UV microscopy (%) DSC' (%)

120°C 0'58 ± 0'01 42 46 125°C 0'52 ± 0'01 48 130°C 0'53 ± 0'01 47 46

aPrima~ crystallinity, corrected for annealing, based on heat of fusion of 209 J g - for polypropylene.

ratio is of the order of 50 ,um. If this ratio is large compared with the spherulite size, the additive will tend to be concentrated in the spherulite boundaries during crystallization, whereas if it is less than the spherulite size there is a greater tendency for most of the additive to be trapped within the growing spherulite. The atactic fraction in polypropylene has a much slower diffusion rate than typical additives, soD/G is of the order ofl ,um or less. On this scale its distribution may be affected by local variations in the spherulite structure. In addition, the atactic material is commonly present at levels of 5% or more and can actually modify the spherulite structure, rather than simply being redistributed. This effect is illustrated in Fig. 7.4. Atactic polypropylene is not visible by uv microscopy but can be rendered visible by staining. To do this it is necessary to attach fluorescent groups to the polymer. The functional group we use is the dimethylaminonaphthylsulphonyl (dansyl) group, which can be covalently bound to the polymer at high temperatures via the sulphonyl azide: 54

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at room temperature. The arrow indicates the spherulite centre.

The concentration of the staining group must be kept as low as possible to minimize alterations in the properties of the polymer - typically 1 % by weight.

Figure 7.5 shows polypropylene spherulites growing from a polymer containing 10% of a fl uorescen tly la belled a tactic fraction. It can be seen that the fluorescent stain is rejected in qualitatively the same way as the low molecular weight additives. More detailed analysis reveals differences. Table 7.3 shows that there is a significant difference between the expected interfacial partition due to rejection from the crystalline part of the polymer and what is actually observed. There is less than the expected amount of rejection because the spherulite growth front is fibrillar and rough on the 111m scale, such that the atactic material is overtaken by the points of the growing fibrils then trapped as the interfibrillar spaces are filled in. Figure 7.6 shows the reverse experiment, in which the isotactic, crystallizable fraction of the polymer is stained and the non-fluorescent atactic material is rejected at the boundaries.

7.5.4 Polymer Blends The use of the sulphonyl azide provides a very convenient and efficient method of binding a range of fluorescent or absorbing groups on to


FIG. 7.5. Polypropylene sample containing 10% of fluorescently labelled atactic material, viewed in fluorescence during crystallization at 1400 C. Bar = 200 11m

(X 100).

hydrocarbon polymers and has many potential applications in the study of mixing phenomena in polymer systems, including polymer blends. We have used UV microscopy to study the morphology of blends of polyolefins with rubbers having bound UV absorbers in their structures.2 Fayt et aT. 55 have used optical microscopy, in combination with selective dyes, to investigate the morphology of blends of polystyrene or PVC with polyolefins. The potential of techniques of this kind remains to be fully exploited.

7.5.5 Morphological Studies on Crystalline Polymers In Section 7.5.3 it was shown that the observed distribution of an absorbing additive around polypropylene spherulites, either quenched during growth or observed in a hot-stage, is adequately represented as the interaction of the kinetics of spherulite growth and of additive diffusion. The effect of this process is to produce an uneven distribution

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256 P. Calven and N C. Billingham

in which there is no additive in the crystalline phase and the additive is non-uniformly distributed in the amorphous material. Consideration of the diffusion coefficients of typical additives at room temperature suggests that this situation cannot persist for very long and that an equilibrium will quickly be established in which the additive is uniformly dispersed throughout the amorphous phase of the polymer. In looking at this phenomenon we found that long periods of annealing do not lead to uniform distribution of additives in polypropylene, even though the annealing times were much longer than those that should have been required to eliminate any concentration gradients. Further, when additives were allowed to diffuse into polypropylene from solution, their distribution was again found to be uneven. We conclude that the additive must be uniformly distributed in the amorphous phase of the polymer but that the amorphous material is not uniformly distributed within the polymer structure. Thus, after sufficiently long annealing times at elevated temperatures, we can regard the distribution of the additive as reflecting the distribution of the crystallinity of the sample, and this allows uv microscopy to be used as a probe of spherulite structure. 56. 57

We have used this approach to show that polypropylene spherulites are more crystalline at their centres than at their boundaries, and that this effect is particularly prominent in purer (octane extracted) samples as shown in Fig. 7.4. This variation results because rejected and entrapped atactic material limits the full development of crystallinity. We can also look at changes that occur within the spherulite during cooling from the crystallization temperatures (l20-140°C) to room temperature. Small crystallinity changes are well known to occur when crystallized polymers are cooled in this way, and they are detectable by scanning calorimetry and X-ray diffraction. In uv microscopy these changes are much more marked, especially in extracted polymer.57 In fluorescence the spherulite centres become darker and the rather uniform spherulite becomes distinctly fibrous. Figure 7.7 shows the apparent crystallinity changes in different parts of a spherulite as the polymer is cooled; these changes are reversible with heating and cooling. Thus, as spherulites are cooled, crystallization continues in those regions that were of too Iowan average molecular weight or too impure to be able to crystallize at the higher temperature. This occurs to a greater extent in the regions that were initially purer, those close to the leading points of the fibrils and the spherulite centre, and these regions reject the additive. Thus the rejection of mobile additives can be used to

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60 80 100 120 140 160 180 200 Temperature (oe)

257

FIG. 7.7. Reversible local crystallinity (%) changes at the centre (<», boundary (0) and mid-radius (0) of a polypropylene spherulite as a function of

temperature.

investigate the secondary crystallization processes in spherulitic polymers.

We have extended these studies to a number of other polymers, some of which show similar density variations, while others do not. The important factors in determining the degree of rejection in any polymer are the concentrations of non-crystallizable impurities, their diffusion coefficients, and the spherulite growth rate, as outlined above. In comparing different polymers, both the growth rates and the diffusion coefficients are dependent on molecular mobility, so their ratio is expected to change much less than their absolute values. Thus we can regard polypropylene as a reasonable model for other polymers. In this case we see strong rejection of polymeric impurities to spherulite boundaries at high growth temperatures, where rates are slow. Where growth rates are faster, we only see clear rejection from spherulitic centres. Since these effects are more evident in the purer, extracted polymer (Fig. 7.4), we will expect to see more significant effects in the purer polymers. Thus we have seen significant crystallinity variations in linear polyethylene, but isotactic polystyrene, which has quite a large atactic content, shows rather uniform spherulites.

For studies in which the additive acts only as a non-reactive stain it can be chosen for experimental convenience. We have found fluorescent additives, such as Uvitex OB, to be particularly suitable since

258 P Calvert and N C. Billingham

comparisons between the fluorescence and UV absorption pictures can help interpretation and reveal artefacts.

7.5.6 Diffusion Rate Measurements An understanding of the diffusion of small molecules in polymers is of great importance both for their application as packaging and barrier materials and for controlling the migration and loss of stabilizing additives.58 Measurement of diffusion coefficients requires the determination of the concentration profile for the diffusant as a function of time. This has been conventionally performed either by the use of multifilm stacks or by diffusing the additive into a rod of material, followed by sectioning and analysis of the sections. Both methods are timeconsuming since diffusion distances of the order of millimetres are used. Additionally, analysis has usually been by use of radiolabelled additives, with all the expense and problems of synthesis that they imply. In a previous review we showed that the diffusion of uv absorbing compounds in solid polymers can easily be monitored by following the progress of the additive into a sample of polymer in the UV

microscope. The polymer may be in the form of a small rod immersed in a solution of the additive in a solvent that does not swell the polymer; for polypropylene suitable solvents are water and glycerol. The rod is sectioned when the additive has penetrated about 100,um, and the concentration profile of the diffusant within the polymer is measured by UV microscopy of the sections. An alternative to solution immersion is to pack a layer of finely powdered solid additive between films of polymer and to clamp the sandwich together for the experimental time. In this case, or if a saturated solution is used, the solubility of the additive in the polymer can also be calculated.

More recently we have found that this approach can be improved by measuring the concentration profile in a single film that has been exposed to a solution of the additive. Figure 7.8 shows the waveform monitor trace resulting from diffusion of a UV absorbing additive into a thin film of polypropylene from one side. The concentration profiles are usually measured from enlarged photographs of the waveform monitor trace. Once the profile is established, the diffusion coefficient of the additive may be determined by fitting the profile to the expected form. 59. 6o Figure 7.9 shows data determined in this way for a strongly UV

absorbing molecule diffusing into polypropylene. The data are all fitted with a single diffusion coefficient of 1·03 X 10-9 cm2 S-I and agreement


FIG. 7.8. Distribution of a uv absorbing additive diffusing into a polypropylene sheet. TV image of UV transmission picture and corresponding waveform

monitor trace.

with experiment is good. This method can only be used with molecules having strong UV absorption or fluorescence, but this is not a severe limitation for polymer stabilizers. Its main advantage is that the concentration profile is measured over a distance of only about 100-200 11m, so the method is much quicker and can be used over a wider range of temperatures than the more conventional macroscopic methods. Data obtained by this method are in good agreement with values measured by Johnson and Westlake61 for the same diffusants.

Although we have mainly used UV absorbers, the same methods can be applied to fluorescent molecules. As an example, we used fluorescence microscopy to monitor the diffusion into polypropylene of an atactic fraction of molecular weight about 8000, rendered visible by covalently bound fluorescent groups as described earlier. Figure 7.10 shows data for diffusion at 130° C for two different times, fitted by diffusion coefficients of around 2 X 10-9 cm2 S-I.

An alternative approach to measuring diffusion coefficients is to begin with a polymer film that contains the dissolved additive uniformly distributed through it. If the film is placed in contact with a liquid that is a good solvent for the additive and non-swelling for the

260

o u

U I

o U

P. Calvert and N. C. Billingham

200 Distance (~m)

300

FIG. 7.9. Experimental data and theoretical curves for diffusion of Aduvex 2412 (2-hydroxy-4-dodecyloxybenzophenone) into polypropylene at 60°C. D =

1'03 X 10-9 em S-I. (1) 22 h. (2) 30 h. (3) 46 h.

polymer, then loss of the additive from the surface is controlled by the rate of diffusion of the solute to the surface. Figure 7.11 shows waveform monitor traces of the concentration profiles in this situation for a typical uv absorber. To a good approximation, the concentration profile in this situation is given by59

o U

U I

o ~

1·0


200 Distance (J.l,m)

261

300

FIG. 7.10. Experimental data and fitted curves for the diffusion of atactic polypropylene into isotactic polymer at 1300 C. (1) D = 2'98 X 10-9 cm2 s -I; 8 h.

(2) D = 1·83 X 10-9 cm2 S-I; 18 h.

262 P. Calvert and N. C. Billingham

FIG. 7.11. UV light intensity profiles for a UV absorber diffusing out of polypropylene film into methanol at 50°C. (a) O. (b) 46 h. (c) 94 h.

C = ~[(exp( -A) cos TTX) - ~(exp( -9A) cos 3TTX)] CO TT 21 3 21

(7.4)

where A = DTT2t14/ 2, D is the diffusion coefficient and I is the film thickness. Computer fitting of this equation to the measured profile from one edge to the centre of the film then allows the diffusion coefficient to be evaluated. Figure 7.12 shows such an analysis for one of the benzophenones at 40° C. The results are again in good agreement with other data and have been described elsewhere.60


7.5.7 Studies of Polymer Oxidation It is well established58 that the oxidative degradation of polyolefins, whether induced thermally or photochemically, leads to loss of useful mechanical properties at levels of oxygen absorption and of chain scission that are much smaller than might be expected. This is taken to imply that oxidation occurs preferentially in localized regions of the polymer, a conclusion that is supported by the observation that an embrittled polymer will usually recover a large proportion of its toughness upon reprocessing. Two questions are relevant to the discussion of the role of morphology in degradation: (l) Are oxidation products, produced during processing, concentrated by rejection during the subsequent crystallization? (2) Does oxidation occur uniformly throughout the amorphous regions of a polymer or is it localized? In attempting to answer these questions we have applied uv microscopy to the study of oxidation in polypropylene.

When partly degraded, polypropylene contains a variety of carbonyl compounds, such as carboxylic acids, ketones and aldehydes, which

o U

U

025

o 50

•

100 Distance (~m)

• o

FIG. 7.12. Experimental points and fitted curves for diffusion of Aduvex UV9 (2-hydroxy-4-methoxybenzophenone) out of polypropiene at 40°C. (a) D =

1·26 X 10- 10 cm2 S-I; 192 h. (b) D = 1-33 X 10- 1 cm2 S-I; 288 h.


absorb uv below 300 nm. However, the absorption is too weak to be observed directly with a xenon source. Accordingly, we have been developing staining methods to enhance the visibility of oxidation in a polymer which is only slightly oxidized. The first approach has been to use reagents that will react with carbonyl groups in a polymer. Two such reagents have been used, 2,4-dinitrophenylhydrazine (DNPH) and dansylhydrazine (l-dimethylaminonaphthalene-5-sulphonylhydrazine, DNSH) which behaves similarly but is fluorescent and so inherently more sensitive. Unfortunately DNSH is a large molecule with little solubility or permeability in the polymer, so only the surface is stained. DNPH is more satisfactory and oxidation can readily be seen at levels corresponding to about one-quarter ofthe induction time in unstabilized polypropylene. The experimental techniques for oxidation and staining of sections have been described elsewhere.62 The DNPH stain is not ideal, in that a significant amount of the oxidation is extracted by the refluxing isopropanol, and it does not react with all types of carbonyl group. Nonetheless it is a very suitable qualitative marker for local oxidation levels by making visible the local concentration of reactive carbonyl groups which are in the amorphous phase but firmly bound to crystal structure. All the work described here was carried out using DNPH as the staining reagent.

We have attempted to use the uv staining technique to confirm the localization of oxidation in degraded and crystallized polymer, and the results have been summarized and discussed.62 Figure 7.13 shows a section of polypropylene that has been oxidized for 12 hours at 1000 C before staining. It can be seen that much material has been extracted from the spherulite boundaries, but there is no evidence for nonuniform staining of the remaining material. Samples crystallized at 1350 C were similarly uniform. Thus with our staining technique it was concluded that the only material that is rejected to the boundaries is that which is also extracted in the staining process. Non-uniform staining is seen in samples that are partly crystallized and quenched, and in some cases there is a wave of rejected material ahead of the growing spherulite which would be expected from the studies of rejection of other impurities. However, this effect may also be partly due to differences in extraction from the slowly crystallized and from the quenched regions.

Given that impurity species do tend to concentrate at spherulite boundaries, it might be expected that preferential oxidation in these regions would be more rapid in slowly crystallized and segregated samples. In looking for such effects we find evidence for distinctly non-


t (

I (

FIG. 7.13. Section of polypropylene, oxidized for 12 h at 100" C in air before crystallization at 1250 C. Stained with DNPH and viewed in UV light.

Bar = 200 11m (X 115).

uniform oxidation, but there is no sign of this preferring the boundaries. We have similarly looked for evidence of local oxidation in samples photo-oxidized after crystallization, without detecting preferential oxidation associated with the spherulite morphology. Thus, although oxidized material is rejected to spherulite boundaries, it apparently does not enhance the subsequent oxidation of the surrounding structure.62

In the absence of morphological effects on oxidation, an alternative source must be found for the uneven oxidation of typical samples. Using very light compression we were able to produce films where the original polymer particles were not merged. Oxidation and staining showed that about 5% of the original particles are oxidizing much faster than the rest, as shown in Fig. 7.14. Also visible in the sections are clusters of black dots, less than l.um in size. These are in patches that

266 P Calvert and N. C. Billingham

FIG. 7.14. Section of a plaque of lightly moulded polypropylene powder. Original powder granules are retained. Oxidized for 2 h in air at 120° C and stained with DNPH. Note the preferential oxidation of some granules.

Bar = 200 pm (X 110).

correspond to the individual particles, and are more evident in the more oxidized particles. Electron microscopy and elemental analysis by EDAX has shown that these particles are residues from the polymerization catalyst, which are apparently able to catalyse the oxidation of the polymer. The reasons for their non-uniform distribution in the original powder are not clear.

7.5.8 Curing and Permeability of Thermosetting Resins In trying to extend the range of applications of uv and fluorescence microscopy, we have looked at thermosetting resins, with the aim of studying structural inhomogeneities in the resin matrix and the penetration of water into the polymer. There have been many studies63- 65

of inhomogeneities in epoxy resins, mostly concentrating on the nodular structures on the scale of 5-100 nm seen in the electron


FIG. 7.15. Section of an epoxy resin (Epon 828) cured with TETA at 1000 C and stained with DNFB. Viewed in UV light. Dark areas show unreacted amine.

Bar = IOO.um (X 130).

microscope. However, thermosetting resins are complex systems and might be subject to other irregularities on a more macroscopic scale. Such irregularities might be expected to arise if the curing reaction is spatially unstable, such that diffusion of one or other component into those regions that first start to cure leads to local regions of high or low cross-link density. Since curing reactions are exothermic, local heating may also produce density fluctuations, and they may also arise from poor mixing of the highly viscous individual components. In attempting to see such inhomogeneities, we used a standard epoxy resin system (Epikote 828, cured with triethylenetetramine) and used a variety of absorbing and fluorescent probes, either mixed with the uncured resin components or allowed to diffuse in from solution after curing. It was expected that any variations in density would be reflected in variations in the solubility and hence the concentration of the staining compounds.66

268 P. Calvert and N. C. Billingham

When uv absorbing compounds were allowed to diffuse into blocks of cured resin from ethanol solution, we found that the diffusion is extremely slow. The inward progress of the stain, as measured by uv microscopy, appeared uniform with a normal diffusion profile and no evidence of density variations within the resin. When the additives were incorporated into the polymer before curing, two types of inhomogeneity were seen. Long streaks of stain visible in some sections were eventually attributed to poor mixing ofthe stain with the resin. In other samples we found halos of stain around the air bubbles formed in the resin during cure. At present we have no explanation for the latter effect but it is clear that the use of non-reacting stains shows no detectable density variations in cured resins. This implies that any variations in density are too small to be detected as variations in additive concentration « 10%) or too small to be resolved in the optical microscope « 1-5 .urn).

In extending this study we used 2,4-dinitrofluorobenzene as a staining reagent. This falls into the category of reactive stains, since it reacts with secondary and primary amino groups to produce strongly uv absorbing centres. This reagent could therefore be used to reveal unreacted amino groups in cured resins. Figure 7.15 shows a UV

micrograph of a resin treated with this reagent after thorough mixing of the components with a glass rod followed by curing at 100° C. The samples show large UV dense regions, indicative oflocal concentrations of unreacted hardener. These concentrations could only be removed by very thorough premixing, either from solution or by using a high shear mixer. Using this technique it is also possible to see variations in the extent of cure arising from temperature variations within samples.66

A third area of study in which UV and fluorescence microscopy can be useful is the penetration of epoxy resins by water.67 There is great interest in this problem, particularly in its effect on interfaces in adhesive joints and in fibre reinforced composites. It would be useful to have a microscopic method for studying water penetration, but it cannot be studied directly by our methods since water neither absorbs nor fluoresces in an accessible wavelength range. Thus we monitor instead the diffusion of a water-soluble UV absorber, 2,4-dinitrophenol, into blocks of fibre reinforced resin; although the phenol will not diffuse as rapidly as the water, its diffusion should be accelerated by the plasticizing effect of the water. Under UV microscopy the reagent diffused smoothly into the unreinforced resin and non-uniform penetration was found only where cracks and bubbles intersected the surface. Samples containing 10% of glass fibre showed no effect of the


FIG. 7.16. Section of Epon 828 containing 5% of glass fibres, cured with TETA then immersed in an aqueous solution of dinitrophenol for 24 h. Viewed in UV light. Note penetration of water along the fibre surfaces. Bar = 300 pm (X47).

glass surface upon the curing reaction, but the fibres provided easy access to water as is shown in the micrograph in Fig. 7.16. The UV microscope thus provides a useful tool for study of water penetration. Its extension to other resins and to adhesive bonding should be fruitful.

ACKNOWLEDGEMENTS

We wish to acknowledge the contribution of past and present members of our research group in developing the techniques described here and for allowing us to quote their unpublished results. Thanks are due particularly to T. G. Ryan, 1. B. Knight, A. Uzuner and Z. Dhanji. We also thank the Science Research Council for the award of a grant to allow the purchase of the uv microscope.


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(1983) 62.

Index

Abbe refractometer, 108 Abbe theory, 46-8 ABS (acrylonitrile-butadiene-

styrene) terpolymers, 65 ABS-modified polycarbonate, 65, 66 Acetal, spherulites, 112, 137, 140, 142 Acrylic cements

embedding in, 26 replication using, 9 thick sections prepared using, 31,

32 Acrylic polymer spheres, refractive

index of, 214, 215 Acrylic sheet, replication using, 11,

177 Additives, investigation of, 52-4, 70 Aduvex 2412, diffusion into

polypropylene, 260 Aduvex UV9, diffusion out of

polypropylene, 263 Anisotropic fibres, refractive index

measurement of, 218 Anisotropic films, refractive index

measurement of, 217-18 Anisotropic materials

meaning of term, 75 refractive index variation in, 80

Anisotropic specimens differential interference contrast

method for, 152, 167

273

Anisotropic specimens-contd. modulation contrast method for,

152, 159-62 Anomalous interference colours, 80 Anti-blocking surface topography,

196, 197 Artefacts, specimen preparation, 1,2 Atomic polarization, 75-7

Babinet compensator, 101-2 Beam damage phenomena, 52-3 Becke line test, 42, 221 Beer-Lambert law, 242 Berek compensator, 102,230 Bertrand lens, 64-5, 98 Biaxial indicatrix, 83-4 Biaxial materials, meaning of term,

75 Birefringence

dispersion of, 104-5 form, 88 orientation effects on, 87, 93-6,

146-8 polarized light produced by, 86 spherulites, 115 strain, 87, 146-7 types of, 87-8

Birefringent films, Abbe refractometer used for, 108

274 Index

Birefringent plates, polarized light passing through, 88-91

Blow-moulded bottles, biaxially orientated, 146, 147

Brace-Kohler compensator, 103 Brewster angle, 85 Brittle materials, sectioning of,

33-4

Cabot carbon test, 53 Calcite crystal, double refraction by,

78, 79 Carbon black, 52, 53 Cauchy relation, 79-80 Circularly polarized light, 81-2

methods of producing, 85-6 spherulitic structures viewed in,

129, 130, 146 Co-extruded products, examination

of,65 Coal, UV microscopy study of, 243-4 Coatings, reflection microscopy

using, 170 Coherence, types of, 188-9 Coherence length, 189 Cold-fracturing technique, 14

disadvantages of, 14 Colours, polarization, 91-3 Comet spherulites, 114, 139 Common light microscopy, 39-40

applications for, 52-60 contaminants detected/identified

by, 54-5, 57 image formation in, 46-51 pigments observed by, 52-4 surfaces examined by, 57-60

Compensation, Senarmont method of, 98, 103-4

Compensators, 100-4 Babinet, 101-2 Berek, 102, 230 Brace-Kohler, 103 Ehringhaus, 102-3,230 interference microscopy, 230-1 quartz wedge, 100-1

Composites, specimen preparation for, 14, 15, 33-4

Contaminants detection/identification of, 54-5,

57 environmental, 57 processing, 55, 57 raw material, 55 sources of, 54-5 see also Cross-contamination

Contrast, definition of, 191 Contrast enhancement methods,

41-2 Copper phthalocyanine, 121 Coverslip method (for interference

microscopy), 207-8, 213-14 Cross-contamination, investigation

of, 70 Crystalline polymers

morphological studies by UV microscopy, 253-8

spherulitic texture of, 65, 67, 112-16

Crystallization, spherulite size affected by, 122, 124

Cut surface technique, 15 Cylindrical specimens, refractive

index measurement of, 215-17

Dark field microscopy, 70-1 Depth reversal phenomenon, 170 Diamond-edged saws, 16,34-5 Dichroism

polarization produced by, 86 UV microscopy use, 243

Differential interference contrast methods, 42, 151-5, 162-83

adjustment of contrast systems, 167-8

anisotropic specimens used, 167 applications of, 173-83, 195 comparison with phase contrast

microscopy, 169 general principles of, 151-5 image interpretation for, 170-2 isotropic specimens used, 162-7 optics used, 162 reflected light microscopy, 169-70 surface microscopy studies, 173-8

Index 275

Differential interference contrast methods-contd.

transmitted light applications, 178-83

Diffraction grating, 47-8 Diffusion rate measurements

fluorescence microscopy used for, 259-62

UV microscopy used for 258-9 1-Dimethylaminonaphthalene-5-

sulphonylhydrazine (DNSH), 264

Dimethylaminonaphthylsulphonyl (dansyl) group, fluorescent labelling with, 250

2,4-Dinitrofluorobenzene, 268 2,4-Dinitrophenol, 268 2,4-Dinitrophenylhydrazine

(DNPH),264 Dispersion

birefringence, 104-5 refractive index, 79-80 see also Relative dispersion; Total

dispersion Double refraction, 77-9

explanation of, 78-9, 80 Double tilt device, 6, 7 Drawn fibres, molecular alignment

of, 94 Drawn films, molecular alignment

of, 95

Ehringhaus compensator, 102-3,230 Electric polarization, 75 Electron microscopy

advantages of, 43-4 comparison with light microscopy,

43-6 Elliptically polarized light, 81-2

methods of producing, 85-6 Embedding techniques, 15, 16,

25-6 acrylic cements used in, 26 epoxy resins used in, 25 ice used in, 26

Embossing, surface topography caused by, 198, 199, 205

Epon 828 resin curing of, 267 glass fibre reinforced, 269

Epoxy resins curing of, 266-8 embedding in, 25, 26, 27 glass fibre reinforced, 269 permeability of, 268-9 thick sections prepared using, 31,

32 Extraordinary (light) ray, 77 Extrudates, surface examination of,

6-7

Fast direction (of polarized light), 88 Fibres

refractive index measurement of, 215-17,218

sectioning of, 27 surface examination of, 12-13

Films anisotropic, refractive index

measurement of, 217-18 anti-blocking additives used in,

196 blocking characteristics of, 196 coated, 186, 203 frictional characteristics of, 196 isotropic, thickness measurement

of, 213-14 sectioning of, 28 surface examination of, 7-11,

173-5 Fluorescence, non-microscopic

applications for polymers, 244-5

Fluorescence microscopy applications to non-polymer

materials, 233, 238-9, 242-3 diffusion rate measurements

using, 259-62 image intensifiers used in, 239 PVC degradation studied by, 246 specimen preparation for, 241 time-lapsed system, 240

Fluorescence microspectrophotometry, 238

276 Index

Foams, specimen preparation for, 15,27

Form birefringence, 88 Fractures

in-service, surface examination of, 6-7

induced, 14 Fresnel-Arago laws, 191 Fringe visibility, definition of, 191

Granular materials, sectioning of, 27

Half-shade plate, 231 Halo effect, phase contrast

microscopy, 63 Heat damage, avoidance of, 53 Hermans orientation factor, 94 HIP (high-impact polystyrene), 65,

179 Hoffmann modulation contrast

methods, 42, 151-62, 167-83 see also Modulation contrast

methods Homogeneous field technique (in

interferometers), 224, 225

Ice, embedding in, 26 Image shading, contrast methods,

171,172 Impurities, rejection during

crystallization, 247-52 Indicatrix. See Biaxial ... ; Optical ... ;

Uniaxial indicatrix Induced-fracture technique, 14 Information processing approach, 50-1 Infrared microscopy, 41 Interference, conditions required,

191 Interference microscopy

accessories fitted to standard equipment, 192

interpretation of surface interferograms, 201-6

mechanical stability requirements for, 192, 209

Interference microscopy-contd. multiple-beam systems, 205, 211-12

image produced by, 206 principle of, 212

nUQlerical aperture effects, 203-4 reflected light methods, 185,

207-12 transmitted light methods, 186,

212-29 two-beam systems, 208-11

disadvantages of, 205 see also Reflected light ... ;

Transmitted light interference microscopy

Interferometry, basic principles of, 186-92

Interphanko interferometers, 217, 222-5

homogeneous field technique used, 224, 225

Isotropic films, thickness measurement of, 213-14

Isotropic materials, meaning of term, 75

Isotropic specimens differential interference contrast

method for, 152, 162-7 modulation contrast method for,

152

Jamin-Lebedeff interferometer, 217, 227-9

principle of, 228

Keith-Padden mechanism (for spherulitic type growth), 117

Kohler illumination, 50, 64, 228, 237 Kohler UV microscopy, 233-4

Laminated products examination of, 65 measurement of refractive index

of, 218 transmitted light interference

image of, 219

Index 277

Lapping techniques, 15-1S manual system used in, 16-17 motor-driven systems used in,

17-1S Lasers

interference using, 190 UV microscopy use of, 239, 240

Light interaction with matter, 73-S0 nature of, 73, 74, lS7

Light microscopy basic principles of, 46-51 comparison with electron

microscopy, 43-6 image formation in, 46-51 practical considerations for, 51-2

Lignin, estimation in wood, 243 Linnik microscope interferometer,

209-10 Liquids, interference microscopy of,

220-2 Lorentz-Lorenz relation, 77

Mach-Zehnder Interphanko system, 222-5

optical basics of, 223 Mach-Zehnder system, 167,223 Magnification considerations, 51-2 Masterbatches, 52 Melinex polyester film, surface

microscopy of, 175 Melt pressing technique, 36

limitations of, 36 Metallising. See Shadow ... ;

Vacuum metallising Michel-Levy chart, 93, 145 Michelson interferometer, 20S Microdensitometry, photographic

image, 241-2 Microtome knives, 20-4

geometry of knife edge for, 23-4 glass, 22 rigidity of, 23 sharpness of knife edge for, 23 steel,21-2 tungsten carbide tipped, 22, 23, 24 types of, 2, 21-2

Microtomes, 19-20 cold-stage, 20, 21, 26 hot-stage, 20

Microvoids, examination of, 70 Mineral oil contamination, 57 Mirau microscope interferometer,

2l0-II Modulation contrast methods, 42,

151-62, 167-S3 adjustment of contrast systems

for, 167-S anisotropic specimens used in,

159-62 applications of, 173-S3, 195 basic principles of, 156-7 comparison with phase contrast

microscopy, 169 general principles of, 151-5 image interpretation for, 170-2 non-ideal specimens used in,

15S-9 optical details, 155-7 reflected light microscopy, 169-70 surface microscopy applications,

173-S transmitted light applications,

17S-S3 Molecular orientation

polarized light affected by, 115-16, 145-S

see also Orientation Moulding tools, surface topography

of, 19S Mouldings, surface examination of,

6-7 Multiphase polymers, interference

microscopy of, 220 Multiple beam interferometer,

2II-12 principle of, 212 vacuum metallising used with, II

Newton's scale of interference colours, 92-3, 163

Nicol prism, S6 Nomarski differential interference

contrast microscope, 162

278 Index

Nonox CI, rejection during crystallization of polypropylene, 248

Nucleation (of spherulites), 119-22 Nucleic acids, UV absorption by,

238-9 Numerical aperture (NA)

definition of, 48-9, 50 interference microscopy, in, 203-4

Nylon fibres, interference fringe image, 216

Nylon 6.6-polypropylene blends, contrast method used, 179, 180

Optical gradient, detection of, 153-4, 167-8

Optical indicatrix, meaning of term, 82

Optical path difference (OPD) measurement of, 99-100, 168,

212-13,224,226,227 relationship to phase difference,

188 spectrophotometric method of

measuring, 105, 107 wedge method of measuring, 107

Ordinary (light) ray, 77 Orientation

birefringence caused by, 87, 93-6, 115-16, 145-8

fluorescence polarization microscopy studies, 246

polarized light affected by, 115-16, 145-8

Oxidative degradation, UV microscopy studies of, 263-6

Particulate additives common-light observations of,

52-4 dark-ground observations of, 70

Particulate contaminants, identification of, 57

Perspex, oriented, polarized light viewing of, 92

Phase contrast methods, 42

Phase contrast microscopy, 60-5 adjustments for, 64-5 advantages of, 64 applications for, 65-7, 70 basic principles of, 60-2 differential interference contrast

method, compared with, 169 halo effect in, 63 modulation contrast method,

compared with, 169 optics involved, 62 shortcomings of, 62-4

Phase detection, 42 Phase difference

definition of, 187 relationship to optical path

difference, 188 Phase objects, polymer specimens

as, 42 Photoelastic effect, 87, 146-7 Pigments

agglomeration of, 54 common-light observations of,

52-4 dark-ground observations of, 70 nucleating action of, 121 streaking of, 54, 56

Pluta interferometer, 226-7, 230 Polarizability, 77 Polarization, mechanis'ms of, 76 Polarization colours, 91-3 Polarized light

elliptically/circularly polarized, 81-2

methods of producing, 85-6 passage through thin birefringent

plates, 88-91 production by

birefringence, 86 reflection, 85-6 selective absorption, 86

Polarized light qualitative microscopy, 111-48

Polarizer, meaning of term, 85 Polarizing microscope, 96-9

analyser in, 97 Bertrand lens in, 98 body slot in, 97-8

Index 279

Polarizing microscope-contd. eyepiece crosswires, 98 light source in, 99 monochromatic light used, 98 polarizer in, 97 rotating analyser in, 98 rotating stage in, 97 strain-free optics in, 98

Polaroid Corporation materials, 86 Polishing techniques, 15-18

manual system used, 16-17 motor-driven systems used, 17-18

Polycarbonate, ABS-modified, 65, 66 Polyester film, surface microscopy

of, 173-4, 175 Polyester-polycarbonate blend,

contrast methods used, 178 Polyester resin, polarized light

viewing of, 106 Poly( ether ether ketone), carbon

fibre filled, specimen preparation for, 17-18, 34

Polyethylene small-angle light-scattering

pattern from, 144 spherulites, 120, 121, 125, 126, 128,

129, 130, 131, 143 surface microscopy of, 173, 174

Poly(ethylene terephthalate) quenching to glassy state, 124 spherulites, 113, 115, 125, 127 surface microscopy of, 173-4, 175

Polymer blends contrast methods used, 178 UV microscopy of, 252-3

Polypropylene diffusion studies for, 259-63 fluorescent labelling of, 250 orientation in, 246 oxidative degradation of, 263-5 rejection of impurities during

crystallization, 247-52 spherulitic forms of, 114, 120, 123,

132-6,202,255,256-7 Poly(propylene oxide), fluorescent

labelling of, 245 Polystyrene, orientation birefringence

in, 148

Polystyrene-poly(vinyl methyl ether) blends, phase separation in, 245

Poly(vinyl alcohol) as replication material, 9

Poly(vinyl chloride) biaxially orientated bottle, 146,

147 contrast method used, 182, 183 embossed foil, 198, 199 fluorescence from, 246 removal of surface coating from,

198,200 Powders

sectioning of, 25-6 surface examination of, 12-13

Printing inks, dark field microscopy for, 69

Processing, spherulites affected by, 136-43

Propylene-ethylene copolymer spherulites, 67 surface microscopy of, 174, 176,

181, 183 PZO Pluta interferometer, 226-7

basic construction of, 226

Quartz wedge compensator, 100-1 Quasi-monochromatic light, 189

~.

Reflected light interference microscopy, 185, 192-212

applications of, 195-201 interpretation of surface

interferograms, 201-6 specimen preparation for, 192-5 systems used, 207-12

Reflected light microscopy applications of, 169-70 contrast methods used, 170

Reflected light multiple beam interferometer, 211-12

Reflection, polarized light produced by, 85-6

Reflection fluorescence microscopy, 237-8

280 Index

Refractive index definition of, 75 dispersion of, 79-80 measurement of,

anisotropic films/fibres, 217-18 liquids, 220-2 phases in solutions, 218-20 spherical/cylindrical specimens,

214-17 Relative dispersion, definition of, 80 Replication techniques, 8-11

acrylic sheet used, 11, 177 advantages of, 195 materials used in, 9 replicating solutions applied, 9-10 thin films used in, 10

Resonance radiation, 74 Roche-Davis interferometer, 229 Row nucleation (of spherulites), 122,

139 Rubber inclusions, refractive index

of, 220 Rubber toughened materials,

examination of, 65 Rubber tyre compound, carbon

black in, 53, 54

Sample preparation. See Specimen preparation

Scanning electron microscopy advantages of, 12, 43-4 comparison with visible light

contrast methods, 175, 178 rough surfaces examined using,

57,175,178 specimen preparation for, 193

Sectioning techniques, 18-35 cold-stage microtomes for, 20, 21,

26 curling of specimens during, 29-30 diamond saws used in, 34-5 hot-stage microtomes for, 20 knife judder effects, 1, 2, 29 microtome knives for, 20-4 microtomes used in, 19-20 rigidity of sample required, 25 speed of cut for, 24

Sectioning techniques-contd. thick section

large area, 31-2 small area, 32

thin section brittle materials, 33-4 composites, 33-4 hints on technique, 28-30 holding of sample for, 25-8

Selective absorption, polarized light produced by, 86

Self-nucleation (of spherulites), 121-2 Senarmont method (of

compensation), 98, 103-4, 230 Shadow metallising, 11-12 Shish-kebab type structure, 122 Slow direction (of polarized light), 88 Small-angle light scattering, 144-5 Smith double-focus interferometer,

229,230 Spatial coherence, 189 Specimen area, 4-5 Specimen preparation, 1-37

factors affecting approach, 4 fluorescence microscopy, 241 initial approach, 3-4 need to segregate operations, 2 reflected light interference

microscopy, 192-5 scanning electron microscopy, 193 time spent on, 3 ultraviolet microscopy, 240-1

Specimen thickness, 18-19 Spectrophotometric method, optical

path difference measured by, 105, 107

Spherical specimens, refractive index measurement of, 214-15

Spherulites appearance in polarized light, 112,

125 birefringence of, 115 characteristics of, 115 crystallization effects on, 122, 124 degree of branching of, 118-19 effects of processing on, 136-43 fine structure of, 113-15 nucleation and growth of, 119-22

Index 281

Spherulites-contd orientation of chains in, 115-16 phase contrast microscopy of, 65,

67 polarized-light microscopy of,

112-16 polypropylene, 114, 120, 123, 132-6,

202, 255, 256-7 types of, 125-32 X-ray diffraction patterns of, 116

Spherulitic crystallization characteristic features of, 112, 113 theory of, 117-24 UV microscopy studies of, 256-7

Sputter coating techniques, 193 Staining techniques, 36-7, 40-1 Strain birefringence, 87, 146-7 Stress whitening, investigation of, 70 Styrene-butadiene-styrene (SBS)

rubbers, interference microscopy of, 220

Su1phonyl azide, fluorescent labelling using, 250, 252

Surface interferometry applications of, 195-201 interpretation of, 201-6

Surface microscopy, 5-11, 57-60, 173-8

extrudates, 6-7 films, 7-11, 173-5 in-service fractures, 6-7 mOUldings, 6-7

Surface roughness, measurement of, 195-8

Surfaces examination of, 57-60 initial study of, 5-6 replication of, 8-11, 195 shadow metallising of, 11-12 types of, 5 vacuum metallising of, 11, 193

Thermosetting resins curing of, 266-8 permeability of, 268-9

Thin sectioning brittle materials, 33-4

Thin sectioning-contd. composites, 33-4 hints on technique, 28-30 holding of specimen for, 25-8

Time-lapsed fluorescence microscopy, 240

Titanium dioxide pigment: 56, 58-9 Tolansky multiple-beam

interferometer, 211-12 Total dispersion, meaning of term,

80 Transmitted light interference

microscopy, 186, 212-29 isotropic film thickness

measurement using, 213-14 lamin-Lebedeff system, 227-9 Mach-Zehnder Interphanko

system, 222-5 PZO Pluta system, 226-7 refractive index measurement of

anisotropic films/fibres, 217-18 liquids, 220-2 phases in sections, 218-20 spherical/cylindrical specimens,

214-17 Roche-Davis instrument, 229 Smith double-focus instrument,

229 Transmitted light measurement,

212-13 Transmitted light microscopy

contrast methods used in, 178-83 image formation in, 46-7

Tungsten carbide tipped (TeT) microtome knives, 22, 23, 24

Ultraviolet microscopy applications to polymers, 245-69 diffusion rate measurements

using, 258-9 first developed, 233-4 impurity rejection during crystall

ization studied by, 247-52 monochromatic radiation sources

used in, 239-40 morphological studies of

crystalline polymers, 253-8

282

Ultraviolet microscopy-contd. non-polymer applications, 242-4 optics of, 235-40 oxidative degradation studied by,

263-6 polymer blends studied by, 252-3 quantitative measurements using,

241-2 scanning microdensitometry of

photomicrographs used, 241-2

specimen preparation for, 240-1 thermosetting resins studied by,

266-9 TV camera used, 239

Uniaxial indicatrix, 82-3 Uniaxial materials, meaning of

term, 75 Uvitex OB, rejection during

crystallization of polypropylene, 247, 248, 249

Vacuum metallising, 11 Van der Kolk test, 42

Index

Video intensification camera (VIC), 239

Voids, investigation of, 14, 70

Watson microscope interferometer, 208-9

disadvantages of, 209 Wedge method, optical path

difference measured by, 107 Welding, effects of, 139, 141 'Wet-and-dry' paper, 16 White-Spruiell orientation factors,

95 Wollaston prisms, 162 Wood, estimation of lignin in, 243 Wright eyepiece, 100

Zeiss Jena Interphako microscope interferometer, 217

Zeiss (Oberkochen) Jamin Lebedeff microscope interferometer, 217

Zeolites, UV microscopy study of, 243 Zero-order beam, 47-8

Applied Polymer Light Microscopy

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