What is Electron Microscopy?
Electron microscopy (EM) is a technique for obtaining high resolution images of biological and non-biological specimens. It is used in biomedical research to investigate the detailed structure of tissues, cells, organelles and macromolecular complexes. The high resolution of EM images results from the use of electrons (which have very short wavelengths) as the source of illuminating radiation. Electron microscopy is used in conjunction with a variety of ancillary techniques (e.g. thin sectioning, immuno-labeling, negative staining) to answer specific questions. EM images provide key information on the structural basis of cell function and of cell disease.
There are two main types of electron microscope – the transmission EM (TEM) and the scanning EM (SEM). The transmission electron microscope is used to view thin specimens (tissue sections, molecules, etc) through which electrons can pass generating a projection image. The TEM is analogous in many ways to the conventional (compound) light microscope. TEM is used, among other things, to image the interior of cells (in thin sections), the structure of protein molecules (contrasted by metal shadowing), the organization of molecules in viruses and cytoskeletal filaments (prepared by the negative staining technique), and the arrangement of protein molecules in cell membranes (by freeze-fracture).
Conventional scanning electron microscopy depends on the emission of secondary electrons from the surface of a specimen. Because of its great depth of focus, a scanning electron microscope is the EM analog of a stereo light microscope. It provides detailed images of the surfaces of cells and whole organisms that are not possible by TEM. It can also be used for particle counting and size determination, and for process control. It is termed a scanning electron microscope because the image is formed by scanning a focused electron beam onto the surface of the specimen in a raster pattern. The interaction of the primary electron beam with the atoms near the surface causes the emission of particles at each point in the raster (e.g., low energy secondary electrons, high energy back scatter electrons, X-rays and even photons). These can be collected with a variety of detectors, and their relative number translated to brightness at each equivalent point on a cathode ray tube. Because the size of the raster at the specimen is much smaller than the viewing screen of the CRT, the final picture is a magnified image of the specimen. Appropriately equipped SEMs (with secondary, backscatter and X-ray detectors) can be used to study the topography and atomic composition of specimens, and also, for example, the surface distribution of immuno-labels.