Introduction to the

Microscope
Jill Ann R. Paranada
 Microscopy and Measurement

• Microscopes – produce an
enlarged image of an
object
– Used to study
organisms, cells, and
cell parts
• magnification
• resolution
– Microscopes vary in
both magnification and
resolution
2
 Microscopy & Resolution
Magnification:
ability to make things appear larger than they are (Image size/Object
size).
determined by multiplying power of both lenses. e.g.Eyepiece 10X
times Objective power (20X, 40X…)
Highest Maximum magnification is around 1000X for compound
microscope.
Resolution:
The fineness of detail that can be distinguished in an image.
Highest Typical Resolution
Optical Microscope ~200 nm
Electron Microscope ~0.1 nm
Circa 1000AD –
The first vision aid
was invented
(inventor unknown)
called a reading
stone. It was a glass sphere that
magnified when laid on top of reading
materials.
Circa 1284 –
Italian, Salvino D'Armate is credited
with inventing the first wearable eye
glasses.
1590 – Two Dutch eye glass makers,
Zaccharias Janssen and son Hans Janssen
experimented with multiple lenses
placed in a tube. The Janssens observed
that viewed objects in front of the tube
appeared greatly enlarged, creating both
the forerunner of the compound
microscope and the telescope.
1665 – English
physicist, Robert
Hooke looked at a
sliver of cork
through a
microscope lens and
noticed some
"pores" or "cells" in
it.
1674 – Anton van Leeuwenhoek built a
simple microscope with only one lens to
examine blood, yeast, insects and many
other tiny objects. Leeuwenhoek was the
first person to describe bacteria, and he
invented new methods for grinding and
polishing microscope lenses that allowed
for curvatures providing magnifications of
up to 270 diameters, the best available
lenses at that time.
18th century – Technical innovations
improved microscopes, leading to
microscopy becoming popular among
scientists. Lenses combining two types
of glass reduced the "chromatic effect"
the disturbing halos resulting from
differences in refraction of light.
1830 – Joseph Jackson Lister reduces
spherical aberration or the "chromatic
effect" by showing that several weak
lenses used together at certain distances
gave good magnification without
blurring the image. This was the
prototype for the compound microscope.
1872 – Ernst Abbe, then research
director of the Zeiss Optical Works,
wrote a mathematical formula called the
"Abbe Sine Condition". His formula
provided calculations that allowed for
the maximum resolution in microscopes
possible.
1903 – Richard
Zsigmondy developed
the ultramicroscope
that could study
objects below the
wavelength of light.
He won the Nobel
Prize in Chemistry in
1925.
1932 – Frits Zernike
invented the phase-
contrast microscope
that allowed for the
study of colorless and
transparent biological
materials for which he
won the Nobel Prize
in Physics in 1953.
1931 – Ernst Ruska co-invented the
electron microscope for which he won the
Nobel Prize in Physics in 1986. An
electron microscope depends on electrons
rather than light to view an object,
electrons are speeded up in a vacuum
until their wavelength is extremely short,
only one hundred-thousandth that of
white light. Electron microscopes make
1931 – Ernst
Ruska

it possible to
view objects
as small as
the diameter
of an atom.
1981 – Gerd Binnig and Heinrich Rohrer
invented the scanning tunneling
microscope that gives three-dimensional
images of objects down to the atomic
level. Binnig and Rohrer won the Nobel
Prize in Physics in 1986. The powerful
scanning tunneling microscope is the
strongest microscope to date.
•Compound Microscope
•Dissection Microscope
•Scanning Electron Microscope (SEM)
•Transmission Electron Microscope (TEM)
Compound microscopes 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.
 Paulownia Wood c.s.
200x

Frog’s blood
1,000x
A dissection microscope 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.
(also called stereo
microscope)
Head of a moth pupa
60x

Sunflower with moth

pupa in the stem
10x
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.
100,000X magnification.
 pigeon blood

cockroach antenna
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.
200,000X magnification.
Cannot be used to view
living specimens.
 bacillus bacteria
dividing

mitochondrion
• Always carry with 2 hands
• Never touch the lenses with your fingers.
• Only use lens paper for cleaning
• Do not force knobs
• Keep objects clear of desk and cords
• When you are finished with your "scope",
rotate the nosepiece so that it's on the low
power objective, roll the stage down to
lowest level, rubber band the cord, then
replace the dust cover.
 Ocular lens
Body Tube

Revolving Nosepiece
Arm
Objective Lens

Stage
Stage
Clips Coarse adjustment knob
Diaphragm
Fine adjustment knob
Light

Base
Ocular lens

magnifies; where you

look through to see the
image of your specimen.
They are usually 10X or
15X power. Our
microscopes have an ocular
lens power of 10x.
 supports the tube and
connects it to the
arm base
 the flat platform
where you place
your slides
stage
 moves stage (or body
tube) up and down

coarse adjustment knob

 small, round knob on
the side of the
microscope used to
fine-tune the focus of
your specimen
fine adjustment knob
after using the coarse
adjustment knob
 the bottom of the
microscope, used for
support

base
 body tube

connects the eyepiece

to the objective
lenses
the part that holds two
or more objective lenses
revolving nosepiece
and can be rotated to
easily change power
 Adds to the magnification
Usually you will find 3 or
4 objective lenses on a
microscope. They almost
objective lens
always consist of 4X, 10X,
40X and 100X
powers. When coupled
with a 10X (most common)
eyepiece lens, we get total
magnifications of 40X (4X
times 10X), 100X , 400X
and 1000X.
The shortest objective lenses
lens is the lowest power, the
longest one is the lens with
the greatest power. Lenses
are color coded.
The high power objective
lenses are retractable (i.e.
40XR). This means that if
they hit a slide, the end of
the lens will objective lenses
push in (spring loaded)
thereby protecting the lens
and the slide.
Stage clips hold the slides in
place. If your microscope
has a mechanical stage, you
will be able to move the
slide around by turning two
knobs. One stage clips
moves it left and right, the
other moves it up and down.
 controls the amount of light
going through the specimen
Many microscopes have a
rotating disk under the
stage. This diaphragm has
different sized holes and is
diaphragm
used to vary the intensity
and size of the cone of light
that is projected upward into
the slide. There is no set rule
regarding which setting to use
for a particular power. Rather,
the setting is a function of the
transparency of the specimen,
the degree of contrastdiaphragm
you desire and the particular
objective lens in use.
makes the specimen
easier to see

light
 How a Microscope Works
1. Use lenses to make small objects
appear larger
2. Compound light microscope: Two
lenses separated by a tube
3. Lenses magnify an object by bending
the light that passes through the lens

Go to
Section:
The proper way to focus a microscope is
to start with the lowest power objective
lens first and while looking from the
side, crank the lens down as close to the
specimen as possible without touching
it. Now, look through the eyepiece lens
and focus upward only until the image
is sharp. If you can't get it in focus,
repeat the process again.
Once the image is sharp with the low
power lens, you should be able to
simply click in the next power lens and
do minor adjustments with the focus
knob. If your microscope has a fine
focus adjustment, turning it a bit should
be all that's necessary. Continue with
subsequent objective lenses and fine
focus each time.
Rotate to 40x objective, locate desired
portion of specimen in the center of the
field. Refocus very carefully so that the
specimen is focused as sharply as
possible. (Do not
alter focus for the
Following steps )
Partially rotate so that 40x and 100x
objectives straddle the specimen.
Place a small drop of oil on the slide in
the center of the lighted area. (Take care
not to dribble on the stage.)
Put the small drop
of oil directly over
the area of the
specimen to be
Examined.
Rotate so that the 100x oil immersion
objective touches the oil and clicks into
place.
Focus only with fine focus. Hopefully,
the specimen will come into focus easily.
Do not change focus dramatically.
Clean up!: When you have finished for
the day, wipe the 100x oil immersion
objective carefully with lens paper to
remove all oil. Wipe oil from the slide
thoroughly with a Kimwipe. Cleanse
stage should any oil have spilled on it.
Recap the immersion oil container
securely, replace in drawer.
Slide # 9

How to Prepare a Slide

1. Place slide on a flat surface.

2. Place a drop of water on the slide. Add the specimen to the
drop of water (at times, you may want to have the specimen
already on the slide before adding the water).
3. Hold the coverslip by its sides and lay its bottom edge on the
slide close to the specimen. Holding the coverslip at a 45°
angle helps.
4. Slowly lower the coverslip so that it spreads the water out. If
you get air bubbles (looking like little black doughnuts),
gently press on the coverslip to move them to the edge. If
there are dry areas under the coverslip, add a little more water
at the edge of the coverslip. Too much water can be dabbed
off with a piece of paper towel