MICROSCOPES 1

MICROSCOPES

“illumination”

Light (visible) Fluorescent U-V Electron

eyepieces

Monocular Binocular

Resolving Power

N.A. affects Wavelength affects

specimen background

Bright field Dark field

MICROSCOPES 2

PARTS & TERMS

Lenses (ocular, objective) Total Magnification

• ocular mag. X objective mag. Condenser Diaphram

MICROSCOPES 3

PARTS & TERMS

Numerical Aperture (N.A.)optical characteristic of a lens

N.A. = i sin

N.A. increases with magnification

MICROSCOPES 4

PARTS & TERMS

Resolving Power (R.P.)• size of the smallest discernable detail• minimum distance between objects so

that they are able to be distinguished as separate (in

nm)

R.P. = wavelength of illumination(2)N.A.

[ RP of human eye ~ 0.2mm = ______ ]

MICROSCOPES 5

Resolving Power (R.P.)

= 650nm, NA = 0.25R.P. = /2NA = _____nm = _____

= 650nm, NA = 1.25R.P. = /2NA = _____nm = _____

= 450nm, NA = 1.25R.P. = /2NA = _____nm = _____

1300 1.3

180 0.180

260 0.260

MICROSCOPES 6

Resolving Power (R.P.)

R.P. = /2NA

Larger or Smaller R.P. = BETTER ?

R.P. improves as N.A. increases or decreases? R.P. improves as increases or decreases?

MICROSCOPES 7

Types of Scopes

MICROSCOPES 8

Types of Scopes

MICROSCOPES 9

Types of Scopes

Compound Light• Bright or Dark field

• Use blue light or blue filter for shorter • R.P. ~ 0.2

MICROSCOPES 11

Types of Scopes

Fluorescent• Dark field compound light microscope• Uses U-V for side illumination of

specimen, fluorescent parts or dyes give off visible light that is viewed

• R.P. ~ 0.2

MICROSCOPES 12

Types of Scopes

UltraViolet (U-V)• Uses U-V as illumination (shorter )• Image recorded then viewed… still or

video camera image(should NOT look directly at U-V!)

• Special lens material (quartz),glass absorbs U-V

• R.P. ~ 0.15

MICROSCOPES 13

Types of Scopes

Electron Microscopes Use electron beam for illumination,

magnets for “lenses”, video to view

Transmission Electron Microscope (TEM)• beam passes thru, view internal structure

Scanning Electron Microscope (SEM)• beam reflects, see external structure, texture

MICROSCOPES 15

Types of Scopes

Transmission Electron Microscope (TEM)

MICROSCOPES 16

Types of Scopes

Transmission Electron Microscope (TEM)• Special specimen preparation (vacuum)• Working magnification:

whole bacteria….. 8,000x - 10,000xthin section, viruses…. 30,000x - 40,000x

• Maximum magnification ~100,000x• R.P. ~0.001 = ______nm

rickettsia herpes simplex

MICROSCOPES 18

Types of Scopes

Scanning Electron Microscope (SEM)

MICROSCOPES 19

Types of Scopes

Scanning Electron Microscope (SEM)• Special specimen preparation

(vacuum & conductor)• Working magnification:

14,000x - 50,000x

• Maximum magnification ~130,000x• R.P. ~0.01 = ______nm

Q fever(rickettsia)

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