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Surface Labeling of Sialic Acid Residues in BHK Cells and Viral Transformants¹

Department of Pediatrics, University of Pennsylvania School of Medicine, and The Joseph Stokes, Jr. Research Institute, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania 19104

Conditions were found for the direct radioactive labeling of glycoproteins on the membranes of baby hamster kidney fibroblasts and their Rous sarcoma virus transformants with negligible loss of viability. Sialic acid residues were specifically detected by oxidation of the cells with periodate followed by reduction with sodium borotritide. Highly specific incorporation of radioactivity was demonstrated into surface membranes isolated as whole structures from these cells. Comparison was made with a known method which labeled galactose residues utilizing galactose oxidase. It was suggested that, when surface labeling procedures are used which are not extremely gentle to the cells, cell fractionation should be performed.

Gel electrophoresis in sodium dodecyl sulfate enabled comparisons to be made of the sialic acid-containing glycoproteins in the surface membranes from the transformed cells and their normal counterpart, and a number of conclusions were drawn on the exposure of different glycosylated macromolecules to the external environment. These included: (a) differences were observed in the molecular weights and number of species of the prominent sialic acid-containing glycoproteins; (b) labeling of isolated membranes showed additional glycoproteins above M.W. 150,000; (c) the method of oxidation prior to labeling yielded different externally oriented glycoproteins in the normal cells, although the transformants showed no difference; and (d) a number of other differences depended on the method of cell harvest prior to labeling.

Transformed cells appeared to adhere more strongly to the plastic surface of the culture flasks and required longer treatments with a variety of reagents than did their normal counterparts to ensure their complete dislodgement.

¹ Supported by American Cancer Society, Grant BC 109, and NIH Grants CA 14037 and CA 14489.

² Permanent address: Department of Biophysics, Weizmann Institute of Science, Rehovot, Israel.

³ To whom requests for reprints should be addressed.

Received 5/ 9/79. Accepted 1/31/80.