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USE of MICROSCOPE

and
EXAMINATION of LIVING MICROORGANISMS

Light microscopy uses visible or ultraviolet light to illuminate an


object.
The light passes through several glass lenses that alter the
path of the light and produce a magnified image of the object.

Observing and Drawing Objects:


Because the light rays from an object cross before
reaching your eye, the image you see through our
light microscopes will be inverted and upside
down.
Sitting on the stage

Viewed through the lens

Magnification
Magnification: the increase of an object's apparent size.
Total magnification is the product of the magnifying powers of
the individual lenses. The magnifying capability of a
microscope is the product of the individual magnifying powers
of the two lenses;
1- Ocular lens (eyepiece) : The lens nearest the eye to magnify
object 10 times (10X)
2- Objective lens: The lens nearest the specimen to magnify
object 4, 10, 40, and 100 times (4X, 10X, 40X, 100X)
Total magnification = ocular x objective
Resolution: is the degree to which the detail in the specimen
is retained in the magnified image. The ability to see in detail is
essential lest everything appears as an unresolved blur.
Magnifying object by using microscope is useful only if detail
can be accurately preserved and observed.

Resolving Power; is the closest spacing between two points at which the points
can still be seen clearly as separate entities. The smaller the
value for resolving power, the smaller the object that can be
seen distinctly.
Resolving Power;
Wavelenght of the light used/2*Numerical aperture (NA), where NA is N*sin.

N: refractive index
: angle between the most divergent light ry gathered by the lense and the
centered of the lens

Types of Microscope
Compound Microscope
These are light illuminated.
The image seen with this type of
microscope is two dimensional.
This microscope is the most
commonly used.
You can view individual cells, even
living ones.
It has high magnification. However,
it has a low resolution.

Dissection or Stereoscope
is light illuminated.
The image that appears is
three dimensional.
It is used for dissection to get a
better look at the larger
specimen.
You cannot see individual cells
because
it
has
a
low
magnification.
earth worm

Confocal Microscope
This microscope uses a laser light.
This light is used because of the
wavelength.
Laser light scan across the
specimen with the aid of scanning
mirrors.
Then image is then placed on a
digital
computer
screen
for
analyzing.

Scanning Electron Microscope (SEM)


SEM use electron illumination.
The image is seen in 3-D.
It has high magnification and
high resolution.
The specimen is coated in gold
and the electrons bounce off to
give you and exterior view of
the specimen.
The pictures are in black and
white.
Eyes of Mosquito

Transmission Electron Microscope (TEM)


TEM is electron illuminated.
This gives a 2-D view.
Thin slices of specimen are
obtained.
The electron beams pass through
this.
It has high magnification and high
resolution.
Plant cell

STAINING TECHNIQUES
Wet mount, hanging drop, Brownian movement

Simple stain:
- Direct Staining
- Negative Staining
Differential stain
Structural or special stainining
Simple staining is used to observe cell morphology, size and
arragement

Experimental Section
1. Wet Mount
2. Simple staining with cyrstal violet
3. Negative staining