Module 2 Flashcards
1 micrometer (μm) =
10^-6 m
1 nanometer (nm) =
10^-9 m
What makes a good microscope
adequate magnifying power
high resolving power
serves your purpose
provides good contrast
two main types of microscopes
light and electron
microscope that use light waves and mirrors
light microscopes
types of light microscopes
simple, compound/complex, bright-field, dark-field, phase contrast, differential interference contrast, fluorescence, confocal, two-photon
- use electron beams as energy source
- has higher magnification and resolving power
- for objects smaller than 0.2 mm in diameter
- in vacuum
- should have mo water when used
electron microscope
- objects under study are darker
- microscopic field is brightly lit
- gross morphology
- can be used stained or unstained
BRIGHT FIELD
- microscopic field is dark
- objects under study are luminous
- for specimens that are:
- invisible in the ordinary light microscope
- cannot be stained by standard methods
- distorted by staining
- has opaque disc that blocks light
DARK FIELD
- principle is based on variations in the refractive indices
(measure of bending or refracting of a beam of light on
entering a denser medium) - not necessary to fix or stain cells
- detailed examination of internal structure
PHASE CONTRAST
- principle is based on variations in the refractive indices
- advantage: no diffraction halo associated with phase contrast
- disadvantage: the three-dimensional appearance may not represent reality
DIFFERENTIAL INTERFERENCE CONTRAST
- makes use of fluorochromes
- visualizes specimens that fluoresce
- detection of immunological reactions
- can be direct (primary antibody fluorochromes) and indirect (secondary antibody is attached to primary antibody)
FLUORESCENCE
- useful for thick specimens like biofilms
- used to visualize structures
CONFOCAL
- useful for examining living cells within intact tissues
- currently limited to advanced clinical and research laboratories
TWO-PHOTON
types of electron microscopes
TRANSMISSION ELECTRON MICROSCOPE (TEM),
SCANNING ELECTRON MICROSCOPE (SEM),
SCANNING TUNNELING MICROSCOPE (STM),
ATOMIC FORCE MICROSCOPE (AFM)
- ultrastructure in thin section of the cells
- can project images in a much higher resolution—up to the
atomic level of thinner objects - examine viruses
- cells should be killed
- 100,000 X and above magnification
TRANSMISSION ELECTRON MICROSCOPE (TEM)
- surface features of viruses and cells
- reveals a 3-D image
SCANNING ELECTRON MICROSCOPE (SEM)
for observing materials like Pure Gold Surface
SCANNING TUNNELING MICROSCOPE (STM)
for observing Nanocellulose
ATOMIC FORCE MICROSCOPE (AFM)
advantages of examining organisms in living or natural state
- observation of unaltered/undistorted characteristics
- cellular processes can be studied
- motility can be observed
- simple to prepare
disadvantages of examining organisms in living or natural state
- refractive index of cells almost similar to that of water
advantages of examining organisms in stained organisms
- provides contrast
- slides can be preserved
- specimens are killed
disadvantages of examining organisms in stained organisms
- more complicated and tedious to prepare
- expensive
Three basic steps in staining microorganisms:
- Smear Preparation
- Fixation
- Staining
