Role of microscopy in
identification of
microorganisms
INTRODUCTION
• The science of investigating small objects using such
an instruments is called MICROSCOPY
• A MICROSCOPE is an optical instrument that uses a
lens or a combination of lenses to see the objects
that are too small for the naked eye
• PRINCIPLE : is to get a magnified image in which
structures may be resolved, which could not be
resolved with the help of an unaided eye
HISTORY
• In 1590’s hans and zacharias jassen of holland created the 1st
compound microscope
• In 1665 robert hooke developed a 1st laboratory compound
microscope
• In 1672 Anthony van leeuwenhock created a simple microscope that
could magnify to about 200x – 300x. He is called the inventor of
microscope
MICROSCOPE PARTS
Ocular lens
Body Tube
Revolving Nosepiece
Arm
Objective Lens
Stage
Stage Clips
Coarse adjustment knob
Diaphragm Fine adjustment knob
Light
Base
WORKING
• OCULAR LENS
(magnifies image) • OBJECTIVE LENS
( gathers light)
• BODY TUBE
( image focuses)
• BENDING LIGHT : the objective convex lens magnifies and
focuses the image inside the body tube and ocular convex
lens of microscope magnifies it
TERMS & DEFINITIONS
• RESOLUTION : power/ability of a lens to separate or
distinguish small objects that are close together
• Shorter wavelength greater resolution
• MAGNIFICATION : increase of an object’s apparent size
Ocular lens
X
Objective lenses
TOTAL
MAGNIFICATION
CLASSIFICATION
• COMPOUND
MICROSCOPE
MICROSCOPE
• ELECTRON
MICROSCOPE
COMPUND MICROSCOPE
• Light illuminated
• 2- dimensional
• High magnification and low resolution
• Measured in nanometers
1. Bright field microscope
2. Dark field microscope
3. Fluorescence microscope
Bright field microscope
• Convenient, relatively inexpensive and affordable
• Contains 2 lens system for magnifying specimens
and uses visible light
• Produces a dark image against brighter background
Dark field microscope
• Uses visible light with
special condenser
diaphragm
• Produces a bright image of the object against a dark
background
Fluorescence microscope
• Exposes specimen to
ultraviolet, violet or
blue light
• Uses powerful mercury
vapour arc lamp
• Employes 3 sets of
filters to alter the light
• Specimens usually objective
stained with
fluorochromes
• Shows a bright image of the object resulting from the
fluorescent light emitted by the specimen
Endothelial cells
Applications of compound microscope
• Bright field microscope are used to view fixed
specimens or live cells
• Dark field microscopy has recently been used
in computer mouse pointing devices, in order to
allow an optical mouse to work on transparent
glass by imaging microscopic flaws and dust on its
surface
• study of crystals and crystal defects, as well as in
the imaging of individual atoms.
• Fluorescence microscope is used to study the
substances like chlorophyll, riboflavin, vitamin A
• Used to study the structures of proteins
• To identify Y chromosome to determine the sex
of an organisms
ELECTRON MICROSCOPE
• Uses beams of electrons to produce images
• High resolutions and high magnification
• Lack color, always shades of black, gray and white
• Measured in micrometers
1. Transmission Electron Microscope [TEM]
2. Scanning Electron Microscope [SEM]
Transmission electron microscope
• Thin slice of specimen are
obtained & stained with
metal
• Electrons scatter when
they pass through
specimen
• Transmitted electrons
( that do not scatters ) are
used to produce image
• Denser region in specimen, scatters more electrons &
appear darker
• High magnification & high resolution
• 2-dimensional view
Mitochondrion
Scanning electron microscope
• The specimen is coated in gold
• Uses electrons reflected from the surface of a
specimen to create image
• 3-dimensional view
• Pictures are in black and white
(a) Ant (b) cockroach’s antennae
Applications of electron microscope
• SEM is used 3D tissue imaging
• Most commonly used for surface characterization
of cells and viruses
• TEM is extensively used in nanotechnology for
semiconductor analysis and production
• Measurement of grain size, crystalline size and
atomic arrangements in materials
• For diagnosis ( based on analysis of diseased
tissue ) in medicine and veterinary medicine
• Medical research including the development and
testing of new drugs
• Direct beam-writing fabrication
• Forensics – finger prints
• Mining (mineral liberation analysis)
THANK YOU
BY:
MOHAMMED ALI
