Experiment 1

An Introduction of Light Microscope

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• The optical microscope, often referred to as
the "light microscope", is a type
of microscope which uses visible light and a
system of lenses to magnify images of small
samples.
• The use of a microscope is to provide a
magnified view of objects that are otherwise
too small to be seen by the naked eye.

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 OBJECTIVES
1. To know the basic construction of light
microscope.
2.To master the correct usage of common light
microscope.

• Remember, microscopes are

expensive scientific
instruments. Handle them
properly and carefully and they
will last for many years!
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 The main construction of
common light microscope
• Laboratory binocular light
microscopes may vary
slightly in design depending
on the manufacturer (e.g.
Nikon, Zeiss, Olympus), but
they comprise of the same
components and similar
principles of operation
apply. 4
 The main construction of
common light microscope
• All modern optical microscopes designed for
viewing samples by transmitted light share the
same basic components of the light path,

The vast majority of

microscopes have the
same 'structural'
components:
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The main construction of
common light microscope

Single lens (simple) Compound microscope

microscope 6
The main construction of common light microscope

• Base
• Supports the
microscope

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The main construction of common light microscope

• Pillar

• The microscope
pillar is the portion
that is mounted to
the base and adds
support to the unit.
It is the main piece
of the microscope
that stands upright,
and is sometimes
used for focusing

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The main construction of common light microscope

• Focus wheels
• Coarse-
focusing knob:
Moves the stage up
and down
(quickly) for
focusing your
image

• Fine-focusing
knob: moves the
stage slightly to
sharpen the image 9
The main construction of common light microscope

Arm
Used to support
the microscope
when carried.
Holds the body
tube, nosepiece
and objective
lenses
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The main construction of common light microscope

• Objective Revolver
or Objective turret
or Revolving nose
piece
• The Nosepiece holds the
objective lenses and can
be turned to increase the
magnification

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The main construction of common light microscope

• Stage
• Supports the
slide/specimen

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The main construction of common light microscope

• Clip
• These clip hold
the
slide/specimen
in place on the
stage.

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The main construction of common light microscope

• Lever to move
stage clip
(control knob)
(Horizontal
specimen position
adjustment )

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The main construction of common light microscope

• Condenser
• A condenser is
a lens that serves
to concentrate
light from the
light source ,
• focus light from
the illumination
source onto the
sample.
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 The main construction of common light microscope

• Diaphragm: a thin opaque structure with an

opening (aperture) at its center.
• The role of the diaphragm is to stop the passage of
light, except for the light passing through
the aperture.

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The main construction of common light microscope

• Diaphragm
• The Diaphragm
controls the amount
of light on the
slide/specimen

Turn to let more light in

or to make dimmer.
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The main construction of common light microscope

• Light source

• Projects light
upwards through
the diaphragm, the
specimen and the
lenses

• Some have lights,

others have
mirrors
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The main construction of common light microscope

• Ocular lens
(eyepiece)

• Magnifies the
specimen image

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The main construction of common light microscope

• Objective
• The Objective Lenses
increase magnification
(usually from 4x to
40x)
• (4x,10x, 40x, 100x)

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 Total Magnification
• The total magnification of a microscope is determined
by multiplying the individual magnifications of the
objective and eyepiece.
• Ocular 10x Objective 40x: 10 x 40 = 400
So the object is 400 times “larger”

Objective Lens have their

magnification written on
them.

Ocular lenses usually magnifies by 10x

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The main construction of common light microscope

• Mechanical components
base, arm, focus wheels, nosepiece, stage
• lighting components
light source, condenser, diaphragm
• Optical Components
eyepiece, objective

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The main construction of common light microscope

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 PROCEDURE
• 1. Carrying a Microscope, and remove the cover.

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 PROCEDURE
• 2. Locate the main parts of light
microscope.
• Make sure it’s on a flat surface.

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 PROCEDURE
• 3. Sit down, and then adjust the seat.

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 PROCEDURE
• 4.Insert the plug into the socket, press the button
to turn on the light, rotate the regulating knob to
make the light intensity proper for you. (Not too
dim, and not too bright!)

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 PROCEDURE
• 5. Set the eye distance on the binocular piece of the
microscope . (There is a ruler between the two
ocular lenses. Make it long or short to suit the
distance of your eyes.)
• Keep both eyes open to reduce eyestrain.

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 PROCEDURE
• 6. Rotate the coarse-focusing
knob to lower the stage down.
First watch the slide directly in
order to know the position of
the specimen, and then put the
slide on the stage and hold it
firmly with the clip (ensuring it
is the right way up, cover slip
facing the objective) , and move
the specimen to the center of
light pore. 29
 PROCEDURE
• 7. Adjust the nosepiece and make 4X lens (low
power magnification) click into working site.

Always observe
using the
LOWEST
POWER
objective first.

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 PROCEDURE
• 8. Rotate the coarse-
focusing knob backward to
raise the stage until you
cannot move it any more.
• Rotate the knob forward
slightly, just a bit, then you
can observe the image
through the ocular lenses.

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 PROCEDURE
• Rotate the fine-focusing knob to make the
image clear.
• Focus using the COARSE ADJUSTMENT
KNOB to bring the object into focus.
• Bring the object into sharp focus by using
the fine adjustment knob.

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• Move the slide so that the image is in the
center of the field of view
• And readjust the light source, condenser
or diaphragm for the clearest image.

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 PROCEDURE
• 9. If you want to see some part more clearly, move
the part to the center of the field of vision. Rotate
the nosepiece to 40X lens (high power
magnification). Rotate fine-focusing knob to make
the image clear.

• Never use coarse-focusing knob for 40X lens !

• Do not allow the objective lens to touch the slide!
Use only the FINE ADJUSTMENT KNOB when
using the HIGH POWER OBJECTIVE.
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• Adjust fine-focusing knob to get clear
image. 35
36
 PROCEDURE
10.Change a slide
• After finishing one slide, if you want to
observe another slide, rotate the nosepiece
to 4X lens, remove the old slide, and replace
the new slide.
• Don’t change the slide under high
magnifient.

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 PROCEDURE
11. After observing,
• Rotate the nosepiece to make all the
objective lenses away from light pore.
• Lower the stage down, and remove the slide.
• Decrease the light intensity to “0”, and turn
off the light. Remove the plug from the
socket ， wrap the cord around the base 。

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 PROCEDURE
11. After observing,
• Return dustcover, always keep the
microscope covered when not in use. Dust
is the number 1 enemy!

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 Image Brightness
• Regardless of the imaging mode utilized in
optical microscopy, image brightness is
governed by the light-gathering power of the
objective, which is a function of numerical
aperture. Just as brightness of the microscope
source illumination is determined by the
square of the condenser working numerical
aperture, brightness of the specimen image is
proportional to the square of the objective
numerical aperture.
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 Numerical Aperture & Resolution
• Numerical aperture as applied to microscope
objectives is a measure of the ability to gather light
and resolve fine specimen detail at a fixed object
distance. The resolution of a microscope objective is
defined as the smallest distance between two points
on a specimen that can still be distinguished as two
separate entities. Resolution is a somewhat subjective
value in microscopy because at high magnification,
an image may appear unsharp but still be resolved to
the maximum ability of the objective. Numerical
aperture determines the resolving power of an
objective, but the total resolution of a microscope
system is also dependent upon the numerical aperture
of the substage condenser. The higher the numerical
aperture of the total system, the better the resolution.41
 CAUTIONS
• 1. Always carry the microscope with two
hands. Hold the arm with one hand, while
support the base with the other hand.
Always carry the microscope in the upright
position so that it will not slip.

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 CAUTIONS
2. Use only lens paper to clean all lenses.
• Use lens paper on all glass parts of the microscope.
• Coarse paper will scratch the lenses. Never allow
your fingers to touch the lenses.

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 CAUTIONS
• 3. If your microscope doesn’t work very
well, please notify you teacher at once.
Never try to repair it by yourself.
Microscope require special repairs to ensure
proper function.
• 4.Never allow the lens to touch the cover
slip or the slide, or wet stains. Corrosive
chemicals may destroy the lens.

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 FINAL CONSIDERATIONS
a. Always look down both eyepieces to produce a single
circular field of view.
b. Move between objectives carefully starting with
lowest (x4 or x10) and then moving to a higher
magnification lens (x40 or x100).
c. Change objectives by rotating the lenses and allowing
them to click into place.
d. Be careful when focusing specimens using a high
power lens – focus with a low power first and then
rotate to high power – move the stage up slowly so not
to crack the slide.
e. Ensure your microscope high power lens is not an oil
immersion lens (it will say so on the lens) otherwise it
will not focus and you may crack the slide.
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• Remember, microscopes are expensive
scientific instruments. Handle them
properly and carefully and they will last
for many years!

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Thank
you !! 47