Electron Microscopy and the Study of the Cell

Electron Microscopy and the Study of the Cell

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Electron Microscopy and the Study of the Cell

Electron Microscopes have revolutionised today’s understanding of the
cell. In 1838 Scheleiden, a botanist theorised that the basic unit of
a plant was a cell, the following year the scientist Schwann came up
with a similar hypothesis this time related to animal cells, their
combined ideas gave us the cell theory, the idea that all living
things were made from similar building blocks, cells. It took 100
years before this idea was confirmed with the invention of the
electron microscope. Prior to this light microscopes, which enable us
to see large cells, were used; but their resolving power was not high
enough to confirm that all living things were made from cells as the
physics of a light microscope only enables it to magnify something by
about 2000x, a resolution of about 0.2 micrometers. In this essay I
will be discussing the uses of electron microscopes, the way in which
they are used and also comparing the electron and light microscopes.
Electron microscopes work on exactly the same principle as their
optical counterparts, but by using electrons as opposed to light. A
stream of electrons is created by a heated filament acting as a
cathode, this stream is then accelerated toward the specimen using a
positive electrical potential, the anode, the specimen lies between
the anode and the cathode. The electrons can be focused into a beam
between metal apertures, using magnetic lenses, as electrons carry a
negative charge, glass lenses are not used as the electrons cannot
pass through glass. The electron beam must pass through a vacuum as
molecules in air would scatter the electrons. This beam is then
focused onto the sample, which is mounted on copper grids for
support. Interactions with the sample create an image, where they are
absorbed or scattered a dark electron dense image is created and where
they are allowed to pass through a light electron dense image is created.
Unlike light lenses, you cannot project this image straight onto the

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eye; the image is focused onto a fluorescent screen, again using
electromagnets to create a picture known as an electron micrograph.

There are 2 types of electron microscope, the scanning electron
microscope and transmission electron microscope. Transmission
electron microscope create a 2D image of the cross-section of a
sample, this microscope type has the highest resolving power.
Scanning electron microscopes give a 3D effect and show surface
detail.

When looking at a specimen under an electron microscope the specimen
must thin enough for electrons to pass through, about as thin as the
film around a soap bubble. There are two ways of preparing a sample
for electron microscopy, negative staining and freezing. Negative
staining involves using an element such as uranium or gold to stain a
sample. An electron micrograph is an electron density map. Elements
such as Carbon, Hydrogen and Oxygen, which make up the largest portion
of most cells, are not electron dense (do not contain many electrons),
so they create a faint electron micrograph. Staining the sample with
elements which are electron dense means that there is a large contrast
and the sample can be seen more clearly. When staining a sample it is
embedded in a resin which becomes hard, when hard it is sliced into
very thin pieces with a glass knife known as a microtome, it is then
treated with a solution of a heavy metal salt like uranyl acetate, and
then dehydrated. The metal ions then surround the exterior of the
specimen; this can cause distortions with the shape of the sample.

To try and prevent this distortion the sample can be frozen with
liquid ethane or liquid nitrogen; because the sample is not dehydrated
it remains the same. The frozen sample is ‘snapped’ into small
slices. Accurate observations are difficult to make with this method
as there are no metal ions present, although the low temperature
decreases damage done by the electrons and allows for a higher
magnification. This method is known as the freeze fracture technique.
Perhaps the most obvious advantage of electron microscopy when
compared to optical microscopy is the clarity and the depth of field
possible to investigate, the wavelength of electrons allows
magnification of up to 500,000x which is huge when compared with the
magnification of just 2000x, that of a light microscope. You might
ask, why use light microscopes at all? Well, as an electron
microscope requires special confinement for its use, as it is affected
by magnetic fields and having large components it is expensive to set
up. The expense does not stop there, the expertise and further
equipment required to prepare specimens means that it is also hugely
expensive to run. The optical light microscope being small, portable,
and with little understanding of light microscopy required to use it
means it is readily available to even the most novice of scientists.
The light microscope allows living cells to be viewed and filmed
whereas the preparation for electron microscopy ensures that nothing
can survive. Natural colours can be viewed with light microscopes and
samples are rarely distorted, false colouring techniques are used on
electron micrographs.

I think these differences mean there is a demand for both microscopes,
although the electron microscope has revolutionised cell biology there
will always be a need for the light microscope.
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