Transcript How to use microscope

General Microbiology Laboratory
Introduction to the oil immersion compound
microscope
• In this lab, you will become familiar with the
use of the microscope (particularly oil
immersion microscopy) and will compare the
relative size and shape of various
microorganisms.
• Most bacteria range in size between 0.5-2.0
micrometers (μm)
• There are three common shapes of bacteria:
the coccus, the bacillus, and the spiral.
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Illustration, cocci
Illustration, bacilli
Illustration, spirochete
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This is the microscope used in our class.
• iris diaphragm lever: moving the
lever to the left increases the light;
moving the lever to the right
decreases the light.
stage control "A": moves the
mechanical stage holding the slide
forward and backward.
stage control "B": moves the
mechanical stage holding the slide
left and right.
coarse focus: turning the knob
away from you raises the stage;
turning the knob towards you lowers
the stage.
fine focus: turning the knob away
from you raises the stage; turning
the knob towards you lowers the
stage.
Proper handling of a microscope. Both hands are used
when carrying this delicate instrument.
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When adjusting the
microscope, the
technologist
observes the
objective carefully
to prevent breaking
the slide and
damaging the
objective lens of the
microscope
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How to use microscope
1.
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4.
Obtain a prepared slide of mixed bacteria. Mount the slide
onto the stage of the microscope.
Start with the lowest power objective in place. Using the
course adjustment knob, move the objective lens to its
lowest point. Look through the ocular and focus upward
with the coarse adjustment until an image comes into view.
Use the fine adjustment to obtain maximum clarity.
Move the slide to a point of interest. Move the next
objective lens into place and adjust the fine focusing knob,
and adjust the iris as necessary. Repeat this step with the
highest power, non-oil lens
Note that as the power of the objective lens increases, the
distance between the objective and the specimen (working
distance) decreases. Also, as magnification increases, the
field of view (visible area) and depth of field/focus (visible
thickness) decrease. Moving the fine adjustment up and
down allows viewing of other areas along the depth of
thickness of the specimen
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Some Concepts to Consider
Definitions and Relationships
• Resolution
• Magnification
• Depth of Focus
• Field of Vision
• Numerical Aperture
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Resolution
• Resolution is the ability to distinguish between
two points.
• The closer the two points, the higher the
resolution.
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Magnification
• Relative enlargement of the
specimen
• The power of magnification is
calculated by multiplying the
power of the eye piece lens by
the power of the objective lens.
• A specimen is magnified as
light passes through the
objective and ocular lens.
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More Concepts…
• Depth of focus - thickness of a specimen
that can be seen in focus at one time;
as magnification the depth of focus .
• Field of vision - the surface area of
view; the area as magnification .
• Numerical aperture (N.A.) – the amount
of light reaching the specimen;
As N.A. resolution .
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Oil immersion
5. To use the oil-immersion lens, move the turret halfway
between the high-power air (non-oil) lens and the oil
lens. Place a drop of immersion oil directly on the slide.
Move the oil-immersion lens into place and adjust the
fine focusing knob. Adjust the iris as necessary. Make
sure that the immersion oil does not get on the air lenses.
Make note of the differences and similarities between the
organisms.
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Resolution can be increased by using immersion oil.
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Cleaning
6. After using the oil lens for a specimen, wipe
the lens with a piece of lens paper. Do not use
anything but lens paper to clean microscope
lenses. Usually, lens-cleaning fluids are not
necessary unless the lens is exceptionally dirty.
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Care and Handling of the Microscope
1. Always use both hands to carry the microscope, one
holding the arm, one under the base.
2. Before each use, examine the microscope carefully
and report any unusual condition or damage.
3. Keep the oculars, objectives, and condenser lens
clean. Use dry lens paper only.
4. At the end of each laboratory period in which the
microscope is used, remove the slide from the stage,
wipe away the oil on the oil-immersion objective, and
place the low-power objective in vertical position.
5. Replace the dust cover, if available, and return the
microscope to its box.
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END OF LECTURE
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