Microscope: P B R.P
Microscope: P B R.P
Microscope: P B R.P
Presented by
R.Parthasarathy
TERMS AND DEFINITIONS
Principle
Microscopy is to get a magnified image, in which structures may be
resolved which could not be resolved with the help of an unaided eye.
Magnification
• It is the ratio of the size of an object seen
under microscope to the actual size
observed with unaided eye.
• The total magnification of microscope
is calculated by multiplying the magnifying
power of the objective lens by that of eye piece.
Resolving power
• It is the ability to differentiate two close points as separate.
• The resolving power of human eye is 0.25 mm
• The light microscope can separate dots that are 0.25µm apart.
• The electron microscope can separate dots that are 0.5nm apart.
TERMS AND DEFINITIONS
Limit of resolution
It is the minimum distance between two points to identify them
separately.
It is calculated by Abbé equation.
Paramecium specimen
Dark field microscope
A bright-field microscope can be adapted as a dark-field microscope by adding a
special disc called a stop to the condenser.
The stop blocks all light from entering the objective lens except peripheral light that
is reflected off the sides of the specimen itself. The resulting image is a brightly
illuminated specimens surrounded by a dark (black) field.
Uses:
This microscope is used to study spirochetes in the exudates form leptospiral or
syphilitic Infections.
Dark field microscope
Paramecium
Treponema vincenti
Uses:
Phase-contrast microscopy is especially useful for studying
microbial motility, studying eukaryotic Cells, determining the
shape of living cells, and detecting bacterial components
such as endospores and Inclusion bodies that contain
poly--hydroxyalkanoates (e.g., poly-hydroxybutyrate),
polymetaphosphate, sulfur, or other substances.
Phase contrast microscope
Macronucleus
Micronucleus
Paramecium
Phase contrast microscope
Rhodospirillum rubrum
Fluorescence microscope
It was developed by Haitinger and coons
A fluorescence microscope differs from an ordinary brightfield microscope in several
respects.
It utilizes a powerful mercury vapor arc lamp for its light source.
A darkfield condenser is usually used in place of the conventional Abbé brightfield
condenser.
It employs three sets of filters to alter the light that passes up through the
instrument to the eye.
Microbiological speciemen that is to be studied must be coated with special
compounds that possess the quality of fluorescence. Such compounds are called
fluorochromes.
AuramineO, acridine orange, and fluorescein are well-known fluorochromes.
Fluorescence microscope
Uses:
Chlamydomonas
Electron microscope(SEM)
The specimen is placed in the vacuum chamber and covered with a thin coat of gold.
The electron beam then scans across the specimen and knocks loose showers of electrons
that are captured by a detector. An image builds line by line, as in a television receiver.
Electrons that strike a sloping surface yield fewer electrons, thereby producing a darker
contrasting spot and a sense of three dimensions.
The resolving power of the conventional SEM is about 10 nm and magnifications with the
SEM are limited to about 20,000x.
Electron microscope(SEM)
Paramecium
SEM
Light Vs Electron microscope
Uses
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