The most commonly used type of electron microscope in biology is called the transmission electron microscope because electrons are transmitted through the specimen to the observer. The transmission electron microscope has essentially the same design as a light microscope, but the lenses, rather than being glass, are electromagnets that bend beams of electrons. An electron gun generates a beam of electrons by heating a thin, V-shaped piece of tungsten wire to 3000°C. A large voltage accelerates the beam down the microscope column, which is under vacuum because the electrons would be slowed and scattered if they collided with air molecules. The magnified image can be viewed on a fluorescent screen that emits light when struck by electrons. While the electron microscope offers great improvements in resolution, electron beams arc potentially highly destructive,and biological material must be subjected to a complex processing schedule before it can be examined. The preparation of cells for electron microscopy is summarized as follows:
A small piece of tissue (approx. 1 mm3) is immersed in glutaraldehyde and osmium tetroxide. These chemicals bind all the component parts of the cells together; the tissue is said to be fixed. It is then washed thoroughly.
The tissue is dehydrated by soaking in acetone or ethanol.
The tissue is embedded in resin which is then baked hard.
Sections (thin slices less than 100 nm thick) are cut with a machine called an ultramicrotome.
The sections are placed on a small copper grid and stained with uranyl acetate and lead citrate. When viewed in the electron microscope, regions that have bound lots of uranium and lead will appear dark because they are a barrier to the electron beam.
The transmission electron microscope produces a detailed image but one that is static,two-dimensional, and highly processed. Often, only a small region of what was once a dynamic, living, three-dimensional cell is revealed. Moreover, the picture revealed is essentially a snapshot taken at the particular instant that the cell was killed. Clearly, such images must be interpreted with great care. Electron microscopes are large and require a skilled operator. Nevertheless, they are the main source of information on the structure of the cell at the nanometer scale, called the ultrastructure.