3H Amino Acid 35S Amino Acid

4C Carbohydrate

3H-Protein, Lipid

35S-Protein, Lipid

14C Glycoprotein Polysaccharide Glycollpid

■ 32P-Protein 32P-Lipid 32P-Nucleic Acid

Figure 1 Biosynthetic labeling of cellular macromolecules. Cells are cultured 2-24 hr in complete cell culture medium or cell culture medium lacking the radiolabeled constituent (e.g., methionine for 35S-methionine). The cells incorporate the radiolabeled precursor into macromolecules and after culture, the cells and culture medium are separated by centrifugation. Radiolabeled macromolecules are extracted from the cell or, if secreted, obtained from the cell culture medium. The radiolabeled macromolecules are then specifically isolated by immunoprecipitation.

radioiodination has identified the molecular properties of B and T cell receptors of antigen, major histocompatibility complex glycoproteins and differentiation antigens. Cell membrane glycoproteins can also be labeled vectorially with 3H-NaBH4 after oxidation of carbohydrate moitiés. In addition, cellular proteins can be labeled by biosynthetic incorporation of 3H, 14C or 35S-labeled amino acid precursors of protein or 3H, 14C-sugars as precursors of carbohydrates. Some proteins can also be labeled by incorporation of 32P catalyzed by various kinases.

After these cells are biosynthetically labeled (see Figure 1), radiolabeled proteins are isolated immuno-chemically, and their structure determined by micro-analytical procedures. In addition, the loss of vec-torially-labeled membrane proteins by cultured cells and the increase in biosynthetically radiolabeled proteins can be taken as a measure of the turnover of cytosolic or membrane proteins.

See also: Antigens, cell surface; Cell surface molecules, immunoprecipitation of; Enzyme labeling of antibodies and antigens; Immunoassays.

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