microscopes 1 “illumination” light (visible) fluorescent u-v electron eyepieces monocular ...
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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
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MICROSCOPES 2
PARTS & TERMS
Lenses (ocular, objective) Total Magnification
• ocular mag. X objective mag. Condenser Diaphram
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MICROSCOPES 3
PARTS & TERMS
Numerical Aperture (N.A.)optical characteristic of a lens
N.A. = i sin
N.A. increases with magnification
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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 = ______ ]
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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
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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?
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MICROSCOPES 7
Types of Scopes
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MICROSCOPES 8
Types of Scopes
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MICROSCOPES 9
Types of Scopes
Compound Light• Bright or Dark field
• Use blue light or blue filter for shorter • R.P. ~ 0.2
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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
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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
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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
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MICROSCOPES 15
Types of Scopes
Transmission Electron Microscope (TEM)
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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
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rickettsia herpes simplex
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MICROSCOPES 18
Types of Scopes
Scanning Electron Microscope (SEM)
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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
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Q fever(rickettsia)
Imageenhancement