This document lists the members of Group One for a microscopy course. It includes 9 students' names and registration numbers. The document then provides an overview of different types of microscopes, including light microscopes, electron microscopes, fluorescence microscopes, and dark field microscopes. It describes the basic setup, working principles, uses and comparisons of these microscopic techniques.

1. GROUP ONE MEMBERS
NAME
1. BRIAN OCHIENG
2. WYCLIFF OMONDI
3. SILUS ALVIN
4. RUTH JEROP
5. MOSES KIMANI
6. SHARON GICHANE
7. SAMUEL KARANJA
8. SUSAN WANGARI
9. SHARON CHERONO
REG NO
H151-01-2035/2021
H151-01-2506/2021
H151-01-2019/2021
H151-01-2028/2021
H151-01-2009/2021
H151-01-2345/2020
H151-01-2454/2021
H151-01-1883/2021
H151-01-2703/2021

2. INTRODUCTION
• Microscopy –is a broad method used to study biological
processes in cells and tissues. It’s a simple, direct technique
for examining the morphology of cells and their organelles.
• Microscope- optical instruments having magnifying lens or
combination of lenses for inspecting objects too small to be
seen or too small to be seen distinctly by naked eye.
• A good microscope should have at least these properties:

3. CONT’
• 1. Good resolution: ability to produce separate images of
closely placed objects so that they can be distinguished as
two separate entities. The resolution power of,
• Resolution depends on refractive index of die medium. Oil
has a higher refractive index than air; hence, use of oil
enhances the resolution power of a microscope.
• 2. Good contrast: improved by staining die specimen. When
the stains bind to the cells, die contrast is increased.

4. CONT’
• 3. Good magnification: This is achieved by use of lenses.
There are two type of concave lenses used:
• Ocular lens with a magnification power of 10x.
• Objective lens-scanning (4x), low power (10x), high power (40x)
and Oil immersion (100x).
• Total magnification of a field is the product of the
magnification of objective lens and ocular lens:
• Scanning field (40x),Low power field (100x), High power
field (400x), Oil immersion field (1000x)

5. 1. LIGHT MICROSCOPE OR BRIGHT-FIELD
• Uses focused light and lenses to magnify specimens which
cannot be seen by naked eyes.
• Forms a dark image against a brighter background, hence
the name.
• Can be in form of simple light microscope (with single
lens)or compound light microscope (with two sets of lenses).
• The parts in a bright-field microscope are divided into three
groups

6. A. MECHANICAL PARTS
• Base: It holds various parts of microscope, such as the light
source, die fine and coarse adjustment knobs.
• C-shaped arm: It holds the microscope, and it connects the
ocular lens 10 to the objective lens.
• Mechanical stage: The arm bears a stage with stage clips
10 hold the slides and the stage control knobs 10 moves the
slide during viewing. It has an aperture at the center that
permit light to reach the object from the bottom

7. B. MAGNIFYING PARTS
• Ocular lens: The arm contains an eyepiece that bears an ocular lens of 10x
magnification power. Microscopes with two eye pieces are called as
binocular microscopes.
• Objective lens: The arm also contains a revolving nose piece that bears
three to five objectives with lenses of differing magnifying power (4x, 10x,
40x and 100x).
• C. ILLUMINATING PARTS
• Condenser: It is mounted beneath the stage which focuses a cone of light on the slide.
• Iris diaphragm: It controls the light that passes through the condenser.
• Light source: It may be a mirror or an electric bulb.
• Fine and coarse adjustment knobs: they sharpen the image

8. WORKING PRINCIPLE
The rays emitted from the light source pass through
the iris diaphragm and fall on the specimen.
The light rays passing through the specimen is
gathered by the objective and a magnified image is
formed.
 This image is further magnified by 1he ocular lens
10 produce the final magnified virtual image.

9. USES OF LIGHT MICROSCOPE
 Combination of staining and light microscopy is used to
identify different kind of bacteria. Gram staining uses
crystal violet to stain Gram +ve bacteria and safranin to
stain Gram –ve bacteria
 Used by mineralogist to in determination of different shape
of crystals by preparing a sample called thin sections.

10. 2. ELECTRON MICROSCOPE
 Is a microscope with a high magnification and resolution
employing electron beams in place of light and using electron
lenses
 Accelerated electrons are source of illumination
 wavelength of electrons can be up to 100000 times shorter
than that of visible light photons. Thus
 the EM has a much better resolving power than a light
microscope
 It can reveal the details of flagella, fimbriae and
intracellular structures of a cell.

11. TYPES OF ELECTRON MICROSCOPE (EM)
Scanning electron microscope-surface a specimen is
scanned by a beam of electrons that are reflected to form an
image. It has a resolution of 7nm or less.
 Transmission electron microscope - an image is derived
from electrons which have passed through the specimen.
The SEM differs from TEM, in producing an image from
electrons emitted by an objects surface rather than from
transmitted electrons

12. THE ELECTRON PATHWAY
Electrons are generated by electron gun, which travels in high speed.
The medium of travel in EM should be a fully vacuum path because in
air path, electrons can get deflected by collisions with air molecules.
Electrons pass through a magnet condenser and then bombard on a
thin sliced specimen mounted on a copper slide.
The specimens scatter the electrons passing through it, and then the
electron beam is focused by magnetic lenses to form an enlarged
visible image of the specimen on a fluorescent screen
A denser region in the specimen scatters more electrons and therefore
appears darker in the image since fewer electrons strike that area of the
screen.

13. MEASURES TO INCREASE THE CONSTRAST OF
EM
• Staining- a solution of heavy metal which binds
to cell structures and makes cell more opaque
• Shadowing - The specimen is coated with a thin
film of platinum or other heavy metals at 45
degrees so that the metal strikes the
microorganism only on one side.

14. PRINCIPLE OF TRANSMISSION ELECTRON MICROSCOPE

15. Comparision Btw Light And Electron
Microscope

16. 3. FLUORESCENCE MICROSCOPE
Is a technique used in light microscopy that allows the
excitation of fluorophores and subsequent detection of
the fluorescent signals
Principle
When fluorescent dyes are exposed 10 ultraviolet rays
(UV) rays, they become excited and are said to
fluorescence.
they convert this invisible, short wavelength rays into
light of longer wavelengths (i.e. visible light).

17. CONT’
The source of light may be a mercury lamp which emits
rays that pass through an excitation filler.
 The excitation filler is so designed that it allows only short
wavelength UV light (about 400 nm, called as the exciting
wavelength of light) to pass through; blocking all other long
wavelength rays.
 The exciting rays then get reflected by a dichromatic
mirror in such a way that they fall on the specimen which is
priory stained by fluorescent dye. Then the specimen is
focused under the microscope.

19. USES OF FLUORESCENCE MICROSCOPE
Imaging structural components of small specimens, such as
cells
Conducting viability studies on cell populations i.e. Are
they alive or dead
Imaging genetic material within a cell (DNA and RNA)
Viewing specific cells within a large population with
techniques .

20. 4. DARK FIELD MICROSCOPE
A dark field microscope is arranges so that the light
source is blocked off, causing light to scatter as it hits
the specimen.
It is similar to ordinary light microscope however the
condenser system is modified so that the specimen is
not illuminated directly as it directs the light
obliquely.

21. PRINCIPLES
The dark field condenser has a central opaque area that
block light from entering the objective lens directly and has
a peripheral annually hollow area which allows the light to
pass through and focus on the specimen obliquely
Only the light which is reflected by the specimen enters the
objective lens whereas the unreflected light does not enter
the objective.
 As a result, the specimen is brightly illuminated, but the
background appears dark.

22. USES OF DARK FIELD MICROSCOPE
• It is useful for demonstration of very thin bacteria not
visible under ordinary illuminations since the reflection of
light makes them appear larger
• It is useful for demonstration of the motility of flagellated
bacteria and protozoa
• It is used to study marine organisms such as algae
• It is used to study mounted cells and tissues
• It is useful in examining external details such as outlines,
edges, grain boundaries and surface defects than internal
structures.

24. REFERENCES
• Arupba Sankar Stastry, Sandhya Bhat K, (2016), ;
Essential of Medical Microbiology 1st Edition , pg. 9-
14.