Cellular structure, enzyme probes

Probes of Cellular Structure. The membrane matrix of animal cells contains, among other compounds, enzymes, proteins, phospholipids, and triglycerides. The rate and extent of release of these compounds from the membrane during selective enzymatic digestion can be used to probe the degree of exposure of these compounds to the surface environment. 

The membrane is a delicate yet sturdy structure which must be studied intact but cannot be isolated without disruption. In essence, membrane isolation involves the removal of all other parts of the cell. The purity of a membrane preparation is usually measured by the relative increase in specific activity of a membrane-boimd marker enzyme in the membrane preparation over the specific activity of the marker enzyme in the homogenate. In cases where specific membrane markers are lacking, enzymatic activities associated exclusively or primarily with other subcellular components must be demonstrated to be absent in order to show the preparation free from cellular contaminants. Such data, combined with phase-contrast and electron microscopic images of the preparation, are usually accepted as adequate criteria for the measurement of purity. One additional technique, at least for membranes with antigenic properties, is preparation of fluorescent antibodies to the isolated membranes and observation of the antibody localization in whole cells (Kabat and Meyer, 1971). The use of fluorescent probes in determining membrane structure and function has been reviewed (Radda, 1975 Azzi, 1975). 

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