Mic 201 Lecture

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Mic 201 Lecture 2 Microscopy
 1. Light Microscopy  Limitations Magnification
and resolution Numerical aperture Oil immersion
lenses  Types of light microscopes Characteristics
and advantages Two dimensional images Bright
field microscope Phase contrast microscope
Dark field microscope Fluorescence microscope  
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Three dimensional images  Differential
Interference Contrast Microscopy (DIC) Atomic
Force Microscope (AFM) Confocal Scanning Laser
Microscope (CSLM)   2. Electron Microscopy 
Transmission Electron Microscope (TEM) Scanning
Electron Microscope (SEM)
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Mic 201 Lecture 2 Microscopy
Resolution 0.2 µm or 200 nm
 1. Light Microscopy  Limitations Magnification
and resolution Numerical aperture Oil immersion
lenses  Types of light microscopes Characteristics
and advantages Two dimensional images Bright
field microscope
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The microscope pictured is referred to as a
compound light microscope. The term light refers
to the method by which light transmits the image
to your eye. Compound deals with the microscope
having more than one lens. Microscope is the
combination of two words "micro" meaning small
and "scope" meaning view. Early microscopes were
called simple because they only had one lens. The
creation of the compound microscope helped to
advance the field of microbiology light years
ahead of where it had been only just a few years
earlier. Modern compound light microscopes, under
optimal conditions, can magnify an object from
1000X to 2000X (times) the specimens original
diameter.
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The compound light microscope
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Staining cells for microscopic observation
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The Gram stain
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   Types of light microscopes Bright field
microscope Phase contrast microscope Dark
field microscope Fluorescence microscope  
Differences in contrast
Differences in refractive indexes
Light reaching specimens from the sides
Specimens that fluoresce
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Photomicrograph of a field of cells of the
bakers yeast Saccharomyces cerevisiae. (a)
Bright-field.
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Photomicrograph of a field of cells of the
bakers yeast Saccharomyces cerevisiae. (b) Phase
contrast.
Photomicrograph of a field of cells of the
bakers yeast Saccharomyces cerevisiae. (c)
Dark-field
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Three dimensional images 
Living structures
Differential Interference Contrast Microscopy
(DIC)
Use of polarized light.
Atomic Force Microscope (AFM)
Based on atomic repulsive forces and different
refractive indexes.
Confocal Scanning Laser Microscope (CSLM)
Laser light source coupled to a light microscope.
2. Electron Microscopy 
For non-living structures
Transmission Electron Microscope (TEM)
Scanning Electron Microscope (SEM)
Internal structures of cells. External features
only.
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Study guide and homework

• What type of microscopes are used to look at
  three-dimensional images?
• What is a Differential Interference Contrast
  Microscopy (DIC)?
• What is polarized light?
• How does an Atomic Force Microscope (AFM) works?
  Does it allow viewing of living specimens?
• What is a Confocal Scanning Laser Microscope
  (CSLM)? What type of samples can be viewed with a
  CSLM?
• How is a three-dimensional image constructed in a
  CSLM?
• Are electron microscopes useful to study living
  organisms?
• What is the difference between a transmission
  electron microscope (TEM) and a scanning electron
  microscope (SEM)?
• What do electron microscopes use instead of
  lenses? And instead of light rays?

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• Which are the advantages and disadvantages of
  electron microscopes?
• How do you compare the resolution of a light
  microscope with the one of an electron
  microscope?
• Which microscope will you use to look at living
  structures?
• Which ones are useful for non-living structures?
• What type of microscope is better adapted to
  study a thick living specimen?
• Which one is useful to look at internal features
  of a cell in detail? And which one is for
  detailed external features?

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Self-evaluation