Electron microscopy.

Veronica Ouyang 12C

Electron microscopy

The development of the electron microscope (EM) has had a significant impact on science research. Invented in the 1930s, the present day version of the EM can magnify up to 500,000 times and has a resolution of about 1nm. In contrast, the light microscope can magnify an object by a maximum of 1500 times and the resolving power is 200nm. That means organelles, which are only blurred images when viewed with a light microscope, can now be studied in great details. Many new structures therefore have been discovered using the EM.

Instead of using light, the EM uses a beam of electrons to resolve objects. The beam, which is produced by a heated filament, can be bent and focused by electromagnetic lenses, in the same way the glass lenses are used in a light microscope. The image is projected into a cathode ray tube, rather than the retina of the eye, to make it visible to the operator. When suitable sections are found, they are photographed to give a permanent record—an electron micrograph.

The reason why the EM has higher resolving power is that the wavelength of the light used in light microscopes is around 500-650nm. This is much longer than which of the electrons is. That means two objects separated by less than 200nm will appear as one object for the light can not pass though, whereas electrons can.

The EM includes two main types—the transmission electron microscope (TEM) and the scanning electron microscope (SEM). The beam of electrons is transmitted though the specimen in the TEMs, thus the specimen must be extremely thin in order to let the electrons pass though.

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The SEMs can be used to study relatively large 3-dimentional objects. Thin sections are not required as the SEM records the electrons that are reflected off the surface of the object but the surface is coated with a thin film of gold. Although the SEM does not have the revolving power of the TEM, it is more versatile and can be used to observe many kinds of intact structures.

Tissues to be observed though the TEM are subjected to five main preparation procedures. These are:

Fixation—preserves the material in a life-like state using substances, which prevent enzyme action.
Dehydration—removes water, allowing the tissue to be penetrated by the embedding medium by using alcohol.
Embedding—cuts the specimen in a suitable medium into very thin sections (about the thickness of the film around a soap bubble). Usually a plastic or epoxy resin is used to keep the specimen rigid.
Sectioning—cuts the specimen into slices of 20-100nm thick, a machine called an ultramicrotome with a glass or diamond knife is used. Thin slices are mounted on copper grids which provide support.
Staining—using compounds containing heavy metals in order to improve contrast. These substances deflect electrons in the beam and produce dark areas on the electron micrograph. On the other hand, light areas are produced when electrons are able to pass though the specimen. A pattern is produced in this way and converted into an image.

Apparently, such procedures pose a major problem. Each of these stages could alter the real appearance of the cells chemically and physically. Such a deviation is known as an artefact. Sometimes it results in the creation of structures that are completely non-existent in the living cell.

Another problem occurs as the EM can only be used to observe dead material. This is because a vacuum is there inside the EM. Air must be removed or otherwise the electrons would be scattered in all directions by the air molecules. Nevertheless no living organism or cells can exist in a vacuum.

In conclusion, the EM has revolutionised the understanding of the structure and function of cells. However, due to the fact that it is very expensive, large, complex and has to be maintained under controlled conditions, e.g. constant temperature, as well as the mentioned drawbacks. The use of the EMs is normally limited to universities and research establishments. Light microscopes still play a major role in basic scientific observations.