Electron Microscopy

The resolving limit of the electron microscope is consequently several orders of magnitude lower than that of the light microscope, and it thus permits the use of far higher effective magnification. The path of electrons through a transmission electron microscope is directed in a manner analogous to the path of light rays through a light microscope. A beam of electrons projected from an electron gun is passed through a series of electromagnetic lenses. The condenser lens collimates the electron beam on the specimen, and an enlarged image is produced by a series of magnifying lenses. The image is rendered visible by allowing it to impinge on a phosphorescent screen. Since electrons can travel through a high vacuum, the entire electron path through the instrument must be evacuated. consequently, specimens must be completely dehydrated prior to examination. Furthermore, very thin specimens (with the thickness of less 100nm or less) can be observed in the conventional electron microscope, since the penetrating power of electrons through the matter is weak. However, the electron beams of an electron microscope with accelerating voltages of a million kV can penetrate intact cells of more than a micrometer thickness. Such instruments are large and costly, and only a few in existence.In a transmission electron microscope, contrast results from the differential scattering of electrons by the specimen, the degree of scattering being a function of a number and mass of atoms that lie in the electron path. Since most of the constituent elements in the biological materials are of low mass, the contrast of these materials is weak. It can be greatly enhanced by staining with the salts of various heavy metals (e.g., lead, tungsten, uranium). These may be either fixed on the specimen (positive staining) or used to increase the electron.