Desktop Microscope User Guide

Version 1.1.0

4Deep inwater imaging

6589 Chebucto Road

Halifax, Nova Scotia B3L 1L9

Canada

tel: (902) 444-4449

info@4-deep.com

www.4-deep.com

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Contents

1 Introduction 3

2 General Safety Information 3

3 What’s in the Box 3

4 The Microscope and its Components 3

4.1 Camera . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

4.2 Sample Stage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

4.3 Point Source . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

5 Microscope Setup 6

5.1 Step 1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

5.2 Step 2 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

5.3 Step 3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

6 Principle of Operation 6

7 Software 7

8 Tips for Optimal Hologram Acquisition 7

9 Care and Maintenance 8

10 Troubleshooting 8

11 Specifications 9

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1 Introduction

Congratulations on your purchase of the Desktop Microscope. Your microscope comes from aline of instruments that all operate under proprietary digital inline holography technology plat-form, developed by 4Deep. With its new and unique method of data acquisition and analysis,this system is designed to optimize your research of the microscopic world.

2 General Safety Information

This instrument is of safety class 1 and has been built and tested according to EN 610101/IEC 1010-1/VDE 0411-1 safety standards for electrical measurement, control and laboratoryequipment.

Attention!

This microscope contains a class IIIB Laser. Up to 100 mW at 405 nm is emitted from the laserhousing itself. If properly assembled, the light emitted from the pinhole should not pose anythreat of long term damage to an exposed eye. There are no user serviceable parts inside thelight source. If problems are encountered regarding the light source, please return it imme-diately for service. Allow only a qualified technician to disassemble the point source. Do notattempt to remedy any problems with the light source on your own!

Attention!

Always make sure that the proper 12 VDC power supply is used with the microscope. Failureto do so could result in critical damage to the microscope.

3 What’s in the Box

The following is the list of parts shipped as part of the Desktop microscope.

1. Desktop microscope, pre-assembled. See the microscope description in the “Microscopeand its Components” section. Follow the cable connection instructions in the “MicroscopeSetup” section.

2. Power supply (12 VDC).

3. Ethernet cable.

4. Sample holder.

4 The Microscope and its Components

The Desktop microscope consists of 3 primary components:

1. Camera: Housed within the top portion of the microscope.

2. Sample Stage: An x-y-z translation stage

3. Point Source: A laser and pinhole, located in the bottom portion of the microscope.

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Figure 1: Main view of the Desktop microscope

4.1 Camera

(a) SSD scale on theopposite side

Figure 2: The camera vertical control knob

The the digital camera included with the microscope is capable of 15 fps at full (2048x2048)resolution. It is mounted to a translation stage which is controlled by the camera vertical controlknob shown in Fig. 2. This alters the Source to Screen Distance (SSD), which changes themagnification and resolution of the microscope:

Increase SSD Magnification Increases Resolution DecreasesDecrease SSD Magnification Decreases Resolution Increases

Figure 3: Microscope connectors

The power supply for the camera is locatedbelow the Ethernet port of the microscope(see Fig. 3). Camera software controls canbe found in the Octopus software package.

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4.2 Sample Stage

The sample stage has two separate controlknobs, one located next to the laser powerswitch which controls the vertical (z) transla-tion stage shown in Fig. 4, and a second setof vertically aligned knobs on the other side ofthe microscope which control the x-y transla-tions of the stage, in Fig. 4(a).

The sample stage has several custom sample holders available, which are designed to holdstandard microscope slides or sample dishes. The magnification of the microscope is depen-dent on the Source to Object Distance (SOD), and the vertical position of the sample stagecontrols this. Warning: Be careful of the point source when moving the vertical sample stageposition; DO NOT touch the point source.

(a) The x-y controls

Figure 4: The z control knob for the sample stage

Increase SOD Magnification DecreasesDecrease SOD Magnification Increases

4.3 Point Source

Located in the lower portion of the microscope (Fig. 5), the point source is made of an IIIB405 nm laser, a 60x objective and a pinhole a similar size to the wavelength of the laser. Thepinhole is on the exposed platinum foil shown, and is extremely fragile. Please refer to the“General Safety Information” at the start of this manual.

Figure 5: Point source assembly

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5 Microscope Setup

The Desktop microscope system is completely assembled and requires only the supplied powercable and CAT5E communication cable to operate immediately.

5.1 Step 1

Plug the 12 VDC camera power cable into the outlet shown in Fig. 3 on page 4. Some modelsof the Desktop microscope will have a separate 6 VDC power cable for the laser; this is tobe connected to the outlet located just below the switch for the laser, shown in Fig. 4 on theprevious page.

5.2 Step 2

Connect the microscope to the computer with Octopus software installed, with a CAT5 or 6Ethernet cable, to the port located just above the power connection, shown in Fig. 3 on page 4.Turn the laser on with the switch shown in Fig. 4 on the previous page.

5.3 Step 3

Open the Octopus program, and go to the camera tab. The program connects to the cameraautomatically and holograms can be saved or reconstructed in real time.

6 Principle of Operation

The Desktop microscope operates on the principles of holography to image a volume in magni-fication. A 405 nm laser is focused on a 500 nm pinhole of the same order of magnitude as thewavelength of the light, which produces a spatially coherent light source as a reference wave.Light scattering from the objects within the media (water) will interfere with the reference waveto produce an interference pattern which contains spatial and phase information of the objectswithin the volume. This interference pattern, the hologram, is recorded by a CCD camera, andreconstructed mathematically to build images of the objects within the volume.

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The basic principle of digital in-line holographic imaging

The reconstructed images can be further analyzed, saved, or assembled into the volume re-constructions based on the application.

7 Software

The holograms acquired by the Desktop microscope camera need to be properly processed inorder to reconstruct the image.

4Deep provides 2 standard software packages, Swordfish, Octopus, and Stingray, which incor-porate our patented fast hologram reconstruction algorithm as well as additional functionality.

Swordfish is a particle counting software capable of detecting, counting and measuring parti-cles in real time with the speed of up to 15 frames per second.

Octopus is a general purpose research-grade software package that can reconstruct hologramsacquired by the microscope camera at the rate of up to 15 frames per second. Octopus providesadvanced 3D visualization and particle detection capabilities.

Stingray is an automated particle recognition and morphology classification software platform.It develops a user defined database from which it learns the type of objects to be detected.

Please refer to the Swordfish, Stingray and Octopus brochures and manuals available at www.4-deep.com for the additional details on software installation and use.

Custom software solutions can be developed by 4Deep based on customer specifications. Con-tact us for details.

8 Tips for Optimal Hologram Acquisition

Image resolution is dependent on the SOD (Source to Object Distance). As this distancedecreases, the resolution and the magnification increase. Maximum resolution is obtainedby placing the sample/object as close as possible to the Point Source (PS). However, keep inmind that as the SOD distance decreases, the field of view becomes smaller.

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The distance between the camera sensor and the point source - Source to Screen Distance(SSD) of the Desktop microscope is adjustable. Contact 4Deep for the development of customconfigurations for special applications.

In order to achieve accurate images, inline holography requires that a reasonable amount ofthe reference wave (unscattered light) reaches the camera sensor. Too many objects betweenthe PS and the camera can reduce the amount of reference wave reaching the camera and willtherefore result in poor reconstructions.

For moving objects, keep the exposure time of the camera as short as possible while stillmaintaining a visible image. Long exposure times will result in blurry or smeared images. Theexposure can be adjusted in the camera control of the Octopus software.

The PS does not produce a large amount of light; therefore background light from externalsources must be reduced in order to obtain optimal images. Excessive background light canwash out the images and produce low contrast reconstructions.

If possible, always obtain a background image (a hologram containing only the light from the PS,without a sample/object causing scatter). A good background image will allow for backgroundsubtraction when reconstructing holograms, producing higher contrast reconstructions whilereducing noise.

9 Care and Maintenance

Your Desktop microscope requires very little maintenance. Keeping the microscope casingclean and free of lint or dust is recommended. Storing the microscope protected from dirt, dustand moisture will ensure a long operational life. Take all measures to prevent contact withthe point source, as any dust or liquids on the pinhole may prevent the proper operationof the microscope.

10 Troubleshooting

Problem: Camera is not detected by the software.

Solution: Make sure the power supply connector is properly connected to the microscope.Make sure the microscope is connected to the Ethernet port on your computer, and that thepower button is on and indicating a current with the light.

Problem: No light from the point source.

Solution: If there is no light coming from the point source and it is properly connected there maybe an alignment problem with the laser, there may be direct damage to the pinhole, or the laserdiode may have burned out. Send the microscope or point source back to the manufacturer forre-alignment.

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11 Specifications

Resolution 0.5 micronPower 5 WattsFootprint 36 cm x 15 cmWeight 4 kgData transfer Gigabit EthernetComputer operating system Windows XP or laterCommunication & power Ethhernet & 12 VDCHologram size from 2048x2048 to 512x512Frame rate from 16 to 50 fps

List of Figures

1 Main view of the Desktop microscope . . . . . . . . . . . . . . . . . . . . . . . . 4

2 The camera vertical control knob . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

3 Microscope connectors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

4 The z control knob for the sample stage . . . . . . . . . . . . . . . . . . . . . . . 5

5 Point source assembly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

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