Compound Light Microscope

Parts

Eyepiece Also known as the ocular lens

Observation tube also known as body tube, connects the eyepiece lens to the
objective lens.

Arm Raises the objective lenses above the stage. Also used for carrying the
microscope.

Objective lenses the second point of magnification. Usually 4X, 10X, 40X AND 100X.

Stage The platform where the slide is placed for viewing. The slide is attached to the
stage by slide clips. It is movable in four directions by using the stage control.
Coarse & Fine focus knobs coarse focus brings the specimen into focus. Fine focus
fine tunes the focus and increases the detail of the specimen.

Lamp the light source for the microscope. Brightness can be adjusted with the
dimmer.

Iris Diaphragm a rotating disk under the stage varies the intensity of light that is
projected upwards into the slide.

Base the bottom of the microscope used for support.

Who Invented the Microscope?

Zacharias Jansen and the first compound microscope. Then, during the 1590's, two
Dutch spectacle makers, Zacharias Jansen and his father Hans started experimenting
with these lenses. They put several lenses in a tube and made a very important
discovery.

Types of Microscopes

Optical Microscopes

Compound Microscopes - Most people immediately recognize a compound

microscope from seeing them in movies or using them in their student days. The original
microscope design was a compound optical, and it's still the most commonly used
today.

A compound microscope works by illuminating the slide from underneath with a light
bulb. The specimen is then magnified by a series of lenses, one near the slide (called
the objective lens) and one near the top (called the eyepiece). This results in a two-
dimensional image that you can adjust depending on the strength of your lenses.

Compound microscope parts vary per design, but are usually pretty standard. This
makes them easy for anyone to learn and use. Other advantages of compound
microscopes are that they can go up to a high magnification and are affordable for
amateurs, students, and scientists. A disadvantage is that they have a lower resolution,
so your image will never be as crisp and sharp as some more advanced types of
microscopes.

Stereo Microscopes - A stereo microscope differs from a compound microscope in a

few key features. The most critical difference you may notice is that it has two
eyepieces instead of one.
The purpose of the stereo microscope is to produce a three-dimensional image, hence
the two eyepieces that send a different image to the right and left eye. The specimen is
usually lit from above, rather than underneath. This makes the stereo microscope ideal
for dissection, inspection, circuit board work, manufacturing, or use with any opaque
specimen.

Stereo microscopes are very easy to use and are fairly inexpensive, making them ideal
for amateurs, professionals, and people in industries that aren't overtly scientific. They
have a low magnification so you cannot see individual cells, which may or may not be
an advantage depending on your needs. Their biggest use is the ability to create three-
dimensional images. More info about stereo microscopes is here.

Confocal Microscopes - Unlike stereo and compound microscopes, the visible light
source comes from a laser. The laser scans the sample with the help of a series of
scanning mirrors, assembles the image in a computer, and displays the image on a
screen. No eyepieces here.

Because the laser can penetrate a sample deeper than light from a bulb, you can create
a three-dimensional image from a selected depth of the specimen. So you can examine
interior structures of a non-opaque specimen, or look at the surface of an opaque
specimen as deep as the laser light can penetrate. This results in highly selective,
detailed images.

As you can imagine, confocal microscopes aren't for the layperson. They cost tens of
thousands of dollars or more, and are used by research scientists such as molecular
biologists.

These days, many optical microscopes send their image to a computer screen rather
than an eyepiece. Then they're often referred to as digital microscopes, but the light
source and interior parts of the microscope are still the same. These have become
much more popular over the years, because it's way easier to look at a large screen
than a tiny eyepiece!

Electron Microscopes

The next category is electron microscopes. Many people have heard of these, but aren't
clear how they actually work. Put simply, an electron microscope scans with electrons
rather than visible light, resulting in a very detailed (and awesome looking) image. This
works because the wavelength of the electrons is much smaller than the wavelength of
light from a bulb or laser, allowing for greater detail when scanning. There are two main
types of electron microscope:Scanning Electron Microscope (SEM) - A SEM sends a
beam of focused electrons to the sample, which bounce off to create a three-
dimensional surface image. With this method, you can create a picture with high
magnification and high resolution, but it will always be an exterior view.

When using a SEM, the sample must be electrically conductive enough so the electrons
actually bounce off it to create the image. Thus specimens are often coated in a thin
layer of gold or other metal.

Transmission Electron Microscope (TEM) - A TEM works by sending the beam of

electrons through a very thin specimen. So rather than scanning over and bouncing off,
the electrons pass through the sample to create a highly detailed two-dimensional
image.

Since the TEM allows for such incredible interior detail, they're often used in medical
research and nanotechnology.

Electron microscopes are very expensive and technical to use. Thus they're for serious
scientists, not students or hobbyists. There are some disadvantages of electron
microscopes, but their contribution to research is invaluable.

Other Types of Microscopes

Most other types of microscopes are less common and used by research scientists.
These include:

Scanning Probe Microscope - These scan the sample with a physical probe.
The tip of the probe raster scans (goes line by line) the specimen and generates
an image with a computer. Unlike an electron microscope, these scan in normal
air rather than a vacuum (or partial vacuum). But the scanning can be slow and
the maximum image size is limited.

Scanning Acoustic Microscope - These use a principle similar to sonar, in that

they employ sound waves to measure the sample. This type of microscope,
called a SAM, is used to find cracks and voids, detect counterfeit materials, test
for failure and reliability, and perform quality control on physical materials. They
can also provide information on the elasticity of cells for biological research.

X-ray Microscope - This one uses electromagnetic radiation in the form of x-

rays to produce images of tiny objects. Unlike an electron microscope, it can be
used to generate an image of living cells. They're useful in biological research
and metallurgy, as the images are highly detailed.
The Ways to Care for this Instrument

Dirt and Debris

Familiarity with the best possible results obtainable with a specific technique and
procedure permits a user to recognize the consequences of dirty optics. The ability to
compare the expected image and the image actually obtained with respect to optimal
sharpness, contrast, and the absence of contamination-dependent visual artifacts
allows the user to immediately recognize when the microscope is dirty or not. If the
image sharpness or contrast is not optimal, then there is a high probability that the
microscope optics are not clean. In order to determine the location of the dirt, proceed
as follows:

Carefully rotate objectives and cameras a small amount within their thread.

Check the slide and cover slip by moving the specimen while focusing initially on
the upper and then the lower surfaces.

Check the condenser while moving it up and down and if applicable, by swiveling
the front lens slightly.

The affected optical surface is identified when a suspected optical component is moved
and the dirt follows this movement. The single exception to this rule is the camera. Dirt
located within the camera (usually on the optical window protecting the image sensor)
will not move when the camera is moved. A macroscopic check for larger dust particles
and scratches on optical surfaces can be carried out using either a magnifying glass
(with a magnification of 3x to 6x) or an eyepiece viewed through the reverse aperture (in
effect, peering through the opposite end of an eyepiece). A soiled objective front lens is
easily identified by examining an evenly lit surface through the rear aperture of the
objective. The internal lens organization produces an enlarged image of the smallest
contaminants present on the external surface of the front lens. The final check should
always involve an assessment of the improvement in image quality.

Optical Surfaces

Concave or convex surfaces (for example, the front lens of dry objectives and
condensers or the eye-lens of some eyepieces) should be distinguished from planar-
parallel or flat surfaces like the front lens of most of the immersion objectives, as well as
condensers, filters, and the protective glass covering camera sensors or the opening
through which light exits. Concave or convex surfaces are cleaned using either the
cotton or the new polyester swabs as described in the next section. Easily accessible
flat surfaces may be similarly cleaned with soft disposable cellulose wipes. Microscope
optics can be composed either of optical glass, quartz, or polymers. The upper surface
of almost all components will be coated with an antireflection layer to minimize stray
light. Some of these coatings may be wipeable or not, due to their softness. Generally,
antireflection coatings are composed of magnesium fluoride and should only be cleaned
with agents free from ammonia and acid. Several alternatives to hazardous solvents
have been found to be effective in microscope cleaning, and a variety of cleaning
agents, as well as cleaning materials, are recommended by different microscopists and
manufacturers. Some optical components are surrounded by black antireflection
surfaces, which are sensitive to organic solvents. The plastic and rubber parts of the
eyepiece will likewise be attacked by organic solvents, including acetone, methylene
chloride, and chloroform

leaning Agents and Procedures

The goal in cleaning microscope optical surfaces is to completely remove dust and dirt
without leaving any residue of the cleaning agent or damaging the surfaces. The
following equipment is required:

Long, thin wooden sticks, preferably of bamboo (obtainable from Chinese

restaurant suppliers) or a comparable, not too flexible material

High purity cotton

Absorbent polyester swabs for cleaning optical components

Soft cosmetic cellulose tissue specifically designed for optical surfaces

(Kimwipes are suitable for lens cleaning, but typical facial tissues contain hard
particulates that are harmful to optical surfaces)

Dust blower or ear wash bulb (laboratory suppliers, pharmacies)

Distilled water

Freshly prepared solution of 5 to 10 drops of a washing-up liquid in 10 milliliters

distilled water

Solvent for the removal of greasy or oily dirt, such as the Optical Cleaning
Solution L (recipe of 85 percent petroleum ether, 15 percent isopropanol), pure
petroleum ether (analytically pure, boiling point less than 44 degrees Celsius) or,
exclusively for cleaning cover slips, pure acetone

Proper Use and Removal of Immersion Oil

Adhering to the following procedures in the use of immersion oil will significantly ease
the task of removing the oil from microscope components before it causes damage. It is
important to recognize that immersion oils are not inert with respect to either optical or
mechanical microscope components, and if left in contact with the instrument, oil will
penetrate into gears and sliding mechanisms and into crevices between lens elements
and their mounting structures with the potential to cause irreversible damage.

The full utilization of the microscope optical system numerical aperture when immersion
objectives are used requires a double oiling technique in which a single drop of
immersion oil is applied to the top lens surface of the substage condenser and another
single drop on the top of the specimen slide. Only a single drop of oil at each specimen-
optical interface can be accommodated without producing contamination that may be
impossible to remove without complete disassembly or factory servicing of the
instrument. The condenser is then raised just to the point that the oil drop contacts the
lower surface of the slide, and the objective front lens is brought into contact with the oil
drop on top of the slide. It should be stressed that the oil immersion technique is only to
be used with a condenser equipped with an immersion-type top lens, and with
immersion objectives.

How to Avoid Contamination

The opening of the binocular tubes should always be protected either with the
eyepieces or with dust covers. If no dust covers from the manufacturer are available,
aluminum foil is a suitable substitute. The best fundamental method to prevent dust
accumulation is to first cover the microscope with two additional plastic bags and then
the dust cover supplied by the manufacturer. The microscope should never be located
in a position where it could be affected by acidic or alkaline vapors, such as in or near a
wet chemical photographic laboratory.

In tropical regions, microscopes frequently experience a buildup of fungus. Although

there are over 100,000 fungus species, two members of the genus Aspergillus are
believed responsible for most lens deterioration. Optimum growth conditions for these
fungi are relatively high temperature and high humidity, but they are more adaptable to
lower humidity levels than most other fungi. Fungal contamination can best be
minimized by reducing the humidity of the room either by air conditioning or by installing
an infrared lamp above the microscope (at a minimum distance of 150 centimeters or 5
feet). Fungi growing on glass surfaces are not attached by roots and can be wiped off,
but unfortunately, residual corrosion marks remain and the original lens performance
cannot be recovered and the lens must be replaced. The only effective means to
avoid fungal damage to optical components is to prevent its growth in the first
place