MICROSCOPES
“illumination”




Light (visible)
Fluorescent
U-V
Electron
Resolving Power
 N.A. affects
 Wavelength affects
MICROSCOPES
eyepieces
 Monocular
 Binocular
specimen background
 Bright field
 Dark field
1
PARTS & TERMS
 Lenses (ocular, objective)
 Total Magnification
• ocular mag. X objective mag.
 Condenser
 Diaphram
MICROSCOPES
2
PARTS & TERMS
 Numerical Aperture (N.A.)
optical characteristic of a lens
N.A. = i sinq
N.A. increases with
magnification
MICROSCOPES
3
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
R.P. = wavelength of illumination
(2)N.A.
MICROSCOPES
[ RP of human eye ~ 0.2mm = ______m ]
4
Resolving Power (R.P.)
l = 650nm, NA = 0.25
1300
1.3 m
R.P. = l/2NA = _____nm
= _____
l = 650nm, NA = 1.25
260
0.260 m
R.P. = l/2NA = _____nm
= _____
l = 450nm, NA = 1.25
180
0.180 m
R.P. = l/2NA = _____nm
= _____
MICROSCOPES
5
Resolving Power (R.P.)
R.P. = l/2NA
Larger or Smaller R.P. = BETTER ?
 R.P. improves as N.A. increases or decreases?
 R.P. improves as l increases or decreases?
MICROSCOPES
6
Types of Scopes
MICROSCOPES
7
Types of Scopes
MICROSCOPES
8
Types of Scopes
Compound Light
• Bright or Dark field
• Use blue light or blue filter for shorter l
• R.P. ~ 0.2m
MICROSCOPES
9
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. ~ 0.2m
MICROSCOPES
11
Types of Scopes
UltraViolet (U-V)
• Uses U-V as illumination (shorter l)
• 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. ~ 0.15m
MICROSCOPES
12
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
13
Types of Scopes
Transmission Electron Microscope (TEM)
MICROSCOPES
15
Types of Scopes
Transmission Electron Microscope (TEM)
• Special specimen preparation (vacuum)
• Working magnification:
whole bacteria…..
thin section, viruses….
8,000x - 10,000x
30,000x - 40,000x
• Maximum magnification ~100,000x
• R.P. ~0.001m = ______nm
MICROSCOPES
16
rickettsia
herpes simplex
Types of Scopes
Scanning Electron Microscope (SEM)
MICROSCOPES
18
Types of Scopes
Scanning Electron Microscope (SEM)
• Special specimen preparation
(vacuum & conductor)
• Working magnification:
14,000x - 50,000x
• Maximum magnification ~130,000x
• R.P. ~0.01m = ______nm
MICROSCOPES
19
Q fever
(rickettsia)
Image
enhancement