Module 2-Microscopy Parts, care and handling.ppt

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1. Module 2
Microscopy: Parts, care and
handling
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2. Learning Objectives
At the end of this module the learner should be able to:
Describe parts of a microscope and their use
Explain proper storage of the microscope
Describe cleaning the microscope
Explain maintenance of microscopes
Discuss troubleshooting of common problems
associated with the use of a microscope
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3. Content Outline
Microscopy
Parts of a microscope
Microscope storage conditions
Equipment required for microscope maintenance
Cleaning solutions and solvent required to clean
optics, stage and body
Materials required for cleaning and its microscope
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4. Microscopy
The scienceof investigating small objects using a
microscope.
Has an essential role for the diagnosis and management
of many infectious disease through examination of
clinical specimen.
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5. Types of microscopes
1.Simple microscope
An ordinary magnifying glass which may have
a magnification of 5x, 10x, 20x or more.
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6. 2. Compound microscope
Has a much higher magnification power than the
simple microscope.
Enable to see objects as small as 0.1 µm or 100 nm.
Uses at least two lenses positioned at different places.
A magnified image of the object produced by first lens
is further enlarged by a second lens
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7. 3. Electron microscope
Further allow us to see objects as small as 0.5 nm in
diameter or roughly 1/200,000
th
the size we can see
with a naked eye, including viruses and cellular
materials.
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8. Compound Microscope
Categorized based on the type of illumination
system used:
Light microscope
Fluorescent microscope
Dark field microscope
Phase contrast microscope
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9. 9
Based on the number of eyepieces, Two types of
compound microscopes available:
Monocular microscopes
•Single eyepiece
•Convenient for use by beginners and for field work
where there is no electricity.
Binocular microscopes
•Two eyepieces
•Recommended where much microscopic work has to
be done.

10. The resolving power of a microscope
The ability of an objective to distinguish the dots
separately and distinctly.
The limit of usable magnification.
•The human eye can separate dots that are 0.25 mm
in diameter.
•A light microscope can separates dots that are
0.25µm apart.
•The electron microscope can separate dots that are
0.5 nm apart.
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11. Parts of a Compound Microscope
Microscope stand
Mechanical adjustment system
Optics
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12. Compound Microscope
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13. Microscope Stand
Tube
Holds the eyepiece and objectives in line and at the
correct distance
Stage
Flat place in which the specimen to be examined is
placed.
In the center there is circular hole that allow the
light form the lamp to pass through
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14. Microscope stand…
Mechanical stage
Enables the slide to be moved in a controlled way,
either along or across the slide.
Sub stage
Located Immediately below the stage
Holds a condenser lens with an iris diagraph and a
holder for light filters and stops.
Foot/Base
This makes sure that the microscope is placed firmly
on the laboratory bench.
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15. The Mechanical Adjustment System
Coarse adjustment
Used to focus using low-power objectives
Controlled by a pair of large knobs positioned one
on each end of the body
Rotation of these knobs moves the tube with its
lenses or, in some microscopes, the stage up or
down rapidly.
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16. Fine adjustment
Used to focus objectives for high-power objectives
Moves the objectives or stage up or down very
slowly.
Controlled by two smaller knobs on each end of the
slide.
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17. Condenser adjustment
Has an adjustment system for its focusing i.e.
opening and closing of its aperture .
Can also be swung aside to remove it or to exchange
it with another.
Focused by rotating a knob to one side of it.
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18. Optics of a Light Microscope
Objectives
The most important parts as the quality and
magnification of the image depend on it.
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19. Description of objectives according to Lens
diameter and equivalent focal length (EFL)
Objectives Description
Diameter Equivalent focal
length
10x 16mm 2/3 inch
40x 4mm 1/6 inch
100x 2mm 1/12 inch
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20. The low power objective: 10x
Used for initial scanning and observation in
most microscopic works.
Used for initial focusing and light adjustment of
the microscope.
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21. The high power objective: 40x
Used for more detailed study as the total
magnification with 10x eyepiece is 400.
Used for the diagnosis of intestinal protozoal
parasites, urine sediments/cells, casts crystals,
and histological sections.
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22. The oil immersion objectives: 100x
Has a very short focal length and working
distance.
The lens rests almost on a microscopic slide
when in use.
Oil is used to increase the numerical aperture
and the resolving power of the objective.
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23. Ocular (Eyepiece)
A lens that magnifies the image formed by the
objectives.
The usual magnification of the ocular is 10x, others are
4x, 6x, 7x, 15x and 20x.
The higher the power, the greater the total
magnification of the microscope.
The lower the power of the eyepiece, however, the
brighter and sharper is the image.
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24. Condenser Lens
A large lens with an iris diaphragm placed below the
stage.
Directs the beam of light from the light source to the
specimen under examination.
Consists of two or three curved lenses so that the light
can pass to the objectives at a sufficiently wide angle.
Raised and lowered beneath the stage to provide light
based on the type of objective used
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25. Iris diaphragm
Controls the amount of light passing through the
specimen under examination.
Located at the bottom of the condenser, under the
lenses, but within the condenser body.
It can be opened or closed as necessary to adjust the
light intensity.
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26. Mirror
Used in the microscope without built in illumination
The illumination system
The modern compound microscope most often has
a built-in illumination system with a controller to
ensure adequate illumination and comfort for the
microscopist.
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27. Routine Use of Basic Microscope: Steps
1. Place the microscope on a firm bench and not
exposed to direct sunlight.
2. Switch on the light.
3. Place the specimen to be examined on the stage.
4. Select the objective to be used.
Its recommended to begin examination with10x objectives.
Once in focus, all the other objectives also will be in focus.
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28. 5. Focus the objectives
Move the objectives carefully downwards using the
coarse adjustment knob and locking at it from the
side until the lens is near the specimen but not
touching it.
Move the objectives slowly upwards, until the image
comes into view and is sharply focused.
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29. 6. Focus the condenser.
Open the iris of the condenser fully and, focus the condenser on the
detail of the light source until the image appears sharp.
7. Adjust the opening of the condenser iris according
to the specimen examined.
Stained smearsthe condenser iris should be opened
more widely giving a well-illuminated image with fine
details.
Unstained specimenthe condenser iris should be
opened in reduced manner to increase the contrast.
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30. 8. Examine the specimen:
For a higher magnification, swing the 40x into place.
Focus the 40x objectives using the fine adjustment.
For the highest magnification, add a drop of
immersion oil to the specimen and swing the 100x oil
immersion objectives in to place. Open the iris fully to
fill the objectives with light.
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31. Care and Handling of the Compound
Microscope
Good working knowledge and proper care of the
microscope are critical to good diagnostic work.
Always use two hands to carry the microscope -one on
the arm and one under the base.
Never carry the microscope upside down, for the
ocular can and will fall out.
Never expose it to sharp knocks, vibrations,
moisture, dust or direct sunlight.
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32. Always turn off the light when not in use.
Use lens paper which is lightly moistened with ethyl
ether/alcohol (80/20 v/v) to clean all lenses.
Always use the proper focusing technique to avoid
ramming the objective lens into a slide -this can break
the objective lens and/or ruin an expensive slide.
Always carefully place the wire out of harm’s way.
Always replace the cover on the microscope when you
put it away
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33. Materials Required for Cleaning a Microscope
Anti-Mold Strips
Applied to prevent mold.
Replace these strips every 3 years. Remove dust attached to
the microscope with a blower.
Lens paper moistened with ethyl/ether alcohol
Applied to four lenses to clean debris and fingerprints before
and after examination.
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34. Microscope Cleaning Process
Cleaning the eye piece
Cleaning the objectives
Cleaning the microscope stage
Cleaning the microscope body
Cleaning the condenser
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35. Cleaning the eye piece
Blow to remove dust before wiping lens
Clean the eyepieces with a cotton swab
moistened with lens cleaning solution
Clean in a circular motion inside out
Wipe the eyepieces dry with lens paper
Repeat cleaning and drying if required
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36. Cleaning the Objectives
Should be cleaned while attached to the microscope
Moisten the lens paper with the cleaning solution.
Wipe gently the objective in a circular motion from the
inside out
Wipe with lens cleaning paper
Should never be removed from the nosepiece
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37. Cleaning the Microscope Stage
Wipe stage using the cleaning solution on a soft
cloth.
Thoroughly dry the stage.
Repeat the above steps, if required
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38. Cleaning the Microscope Body
Unplug the microscope from the power source
Moisten the cotton pad with a mild cleaning
agent
Wipe the microscope body to remove dust, dirt
and oil
Repeat steps1–3, if required
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39. Cleaning the Condenser
Unplug the microscope from the power source
Clean the condenser lens and auxiliary lens using
lint-free cotton swabs moistened with lens
cleaning solution
Wipe with dry swabs
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40. Maintenance of the microscope:
Replacing the Microscope Bulb
Unplug the microscope from the power source
Find the location of the bulb
Follow manufacturer’s instructions to remove the bulb
Use tissue paper to remove the bulb from the
microscope
Replace the bulb by holding it with lens paper or an
appropriate device
NB: Never touch the bulb with your fingers.
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41. Microscope Repair
Never disassemble the microscope
Optics: eyepieces and objectives
Mechanics: stage and focus adjustments
Repair of these items requires a service engineer
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42. Troubleshooting
1. The brightness of the viewing field is poor
Problem Solution
The condenser is too
low.
Raise the condenser to
correct its position.
The condenser iris
diaphragm is closed.
Open the diaphragm
properly.
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43. 2. Dark shadows in the field which move as
you turn around the eyepiece.
Problem Solution
The surface of the
eyepiece has scratches.
Replace the eyepiece.
The eyepiece is dirty.Clean the eyepiece.
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44. 3. The image with the high power objective is
not clear.
Problem Solution
The slide is upside down.Turn the slide over.
There is an air bubble in
the oil.
Move 100x lens quickly
from side to side.
There is dirt on the
objective.
Clean the lens.
The oil is too sticky.Use thinner immersion oil
or specified immersion oil.
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45. 4. The image is not clear with the low power
objective.
Problem Solution
There is oil on the lens.Clean the lens.
There is a layer of dust on
the upper surface of the
objective.
Clean the lens.
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46. If the field is still dim and cloudy, consider
the following possible causes:
Massive growth of fungus on the lenses or prisms
due to storage in a high humidity environment
Penetration of immersion oil between the lenses of
the objective through damaged lens cement
A damaged objective (due to careless focusing,
dropping, rough changing of slides)
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47. Frequently-encountered operational errors
include the following:
Focusing the first slide using the 100x without passing
through a 10x objective.
Changing slides from under the immersion objective
without turning it away first.
Wiping lenses without first blowing away dust and
sand.
Cleaning lenses or other parts with xylene.
Using cedar wood oil, liquid paraffin, or xylene-diluted
oil instead of pure synthetic immersion oil.
Keeping the microscope in a confined space.
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48. Log Book
It should be maintained to enter problems
encountered in the operation of the microscope,
maintenance schedule, repairs done on the
microscope and availability of spares like bulbs,
fuses, anti-mold strips etc.
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49. Thank You!!
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