Laboratory 2. Microscope

HUMAN ANATOMY AND PHYSIOLOGY WITH PATHOPHYSIOLOGY (LABORATORY)
COMPOUND MICROSCOPE
DEFINITION OF TERMS
➢ Magnification
o The ratio of the apparent size of an
object as seen through the microscope &
the actual size of the object
➢ Resolution/Resolving Power
o The ability of the lens to clearly separate
or distinguish two points of two lines
individually in the image.
o It is determined by the shortest
wavelength of visible light & maximum
numerical aperture - Two lenses
➢ Numerical Aperture - Better magnification than a simple microscope
o A measurement of the ability of the - Bright field microscopes (the specimen is lit from
condenser and the objective lens to underneath, and they can be binocular or
gather light. monocular) provide a magnification of 1,000
Bright field Compound Microscope times, which is considered to be high, although
Objective Lens Magnification Aperture the resolution is low.
Scanner 4X 0.10 - Allows users to take a close look at objects too
LPO 10X 0.25 small to be seen with the naked eye, including
HPO 40X 0.65 individual cells.
OIO 100X 1.25
Bright field microscopes
➢ Focal length
o Thickness of the object that maybe seen
at one time under focus.
o Distance between the center of a lens or
curved mirror and its focus
➢ Working distance
o Distance between the front lens of the
objective lens & the top of the cover
glass when the specimen is in focus.
➢ Parfocal
o Refers to quality of the objectives & ➢ Stained tissue is examined with ordinary light
eyepiece where practically no change in passing through the preparation.
focus has to be made when objective is ➢ The microscope includes an optical system and
substituted for another. mechanisms to move and focus the specimen.
➢ Refractive Index
o Bending of light rays away from the Fluorescence Microscopy
objective lens when light passes from
the glass of the microscope slide to the
air.
o The lower the refractive index of an
object, the better its magnification
SIMPLE MICROSCOPE
- First microscope.
- Created in the 17th century by Antony van
Leeuwenhoek, who combined a convex lens
➢ Tissue sections are usually irradiated with
with a holder for specimens.
ultraviolet (UV) light and the emission is in the
- 200 – 300x Magnification
visible portion of the spectrum.
- Not used often today because the introduction
➢ Fluorescent substances appear bright on a dark
of a second lens led to the more powerful
background
compound microscope
Phase Contrast Microscopy TRANSMISSION ELECTRON MICROSCOPE (TEM)
➢ uses a lens system that produces visible images

from transparent objects and, importantly, can - Transmission electron microscope (TEM) uses
be used with living, cultured cells. electrons in creating a magnified image
➢ it is ideal that the specimen here is not stained - Uses a slide preparation to obtain a 2D view of
because the light is used to provide 3D view of specimens, so it's more suited for viewing
the specimen objects with some degree of transparency.
Polarizing Microscopy - Offers a high degree of both magnification and
resolution
➢ Allows the recognition of stained or unstained

structures made of highly organized subunits.
➢ Tissue structures containing oriented
macromolecules are located between the two
polarizing filters, they appear as bright structures
against a dark background.
➢ It uses polarizing filters to filter out specific
wavelength of light to provide contrasting color.
➢ To view crystalline structures (kidneys and gout)
SCANNING ELECTRON MICROSCOPE (SEM)
- Most powerful magnification

- Uses electrons rather than light for image
formation
MECHANICAL PARTS
- used to support and adjust the parts
- Samples are scanned in vacuum or near vacuum
conditions, so they must be specially prepared by
first undergoing dehydration and then being • Base – bottom post portion that supports the entire
coated with a thin layer of a conducive material, microscope, provides stability and support for the
such as gold. microscope when it is upright. The base also
- After the item is prepared and placed in the typically holds the illuminator, or light source.
chamber, the SEM produces a 3D, black and • Pillar – part above the base that supports the other
white image on a computer screen parts
• Inclination joint – allows for tilting for the
convenience of the user
• Arm – connects the eyepiece tube to the base,
curve, or slanted part which is held while carrying
the microscope
• Stage and stage clip – is a platform for the slides,
which hold the specimen. The stage typically has a
stage clip on either side to hold the slide firmly in
place. Some microscopes have a mechanical stage,
with adjustment knobs that allow for more precise

positioning of slides.
• Body tube – attached to the arm and bears the ✓ Always carry with 2 hands.
lenses ✓ Never touch the lenses with your fingers.
• Draw tube – cylindrical structure on the top of the ✓ Only use lens paper for cleaning.
body tube that holds the ocular lenses. ✓ Keep objects clear of desk and cords.
• Revolving nosepiece – contains the objective ✓ When you are finished with your "scope", rotate
lenses. Microscope users can rotate this part to the nosepiece so that it's on the low power
switch between the objective lenses and adjust the objective, roll the stage down to lowest level,
magnification power rubber band the cord, then replace the dust
• Dust shield – lies atop the nose piece and keeps dust cover.
from settling on the objectives
• Coarse Adjustment Knob – moves stage (or body
tube) up and down, used to focus the image on the
microscope under low or medium power
• Fine Adjustment Knob – small knob, used to fine
tune the focus of your specimen after using the
coarse adjustment knob
MAGNIFYING PARTS
- used to enlarge the specimen
• Ocular/ Eyepiece – contains the ocular lens, which

the user looks through to see the magnified
specimen, further magnify the image produced by
the objective lenses by 5x to 15x
• Objective lenses – combine with the eyepiece lens
to increase magnification levels. Microscopes
generally feature three or four objective lenses,
with magnification levels ranging 4x to 100x.
ILLUMINATING PARTS
- used to provide light
• Illuminator or Electric Lamp – microscopes require

a light source for viewing. This can come in the form
of a built-in, low-voltage illuminator light, or a
mirror that reflects an external light source like
sunlight.
• Mirror – has a concave and plane surface to gather
and direct light in order to illuminate the object.
• Condenser – focus the light onto the specimen,
concentrating light rays on the specimen
• Iris Diaphragm – controls the amount of light
reaching the specimen and regulates the light
necessary to obtain a clear view of the object
1. 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.
2. 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.
3. 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
4. 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.

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HUMAN ANATOMY AND PHYSIOLOGY WITH PATHOPHYSIOLOGY (LABORATORY)

Phase Contrast Microscopy TRANSMISSION ELECTRON MICROSCOPE (TEM)

➢ uses a lens system that produces visible images

➢ Allows the recognition of stained or unstained

SCANNING ELECTRON MICROSCOPE (SEM)

- Most powerful magnification

with adjustment knobs that allow for more precise

• Ocular/ Eyepiece – contains the ocular lens, which

• Illuminator or Electric Lamp – microscopes require

1. The proper way to focus a microscope is to start

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