DEVELOPMENT, CHARACTERIZATION, AND APPLICATION OF MONOCLONAL-ANTIBODIES TO THE NATIVE AND SYNTHETIC BETA-COOH-TERMINAL PORTION OF HUMAN CHORIONIC-GONADOTROPIN (HCG) THAT DISTINGUISH BETWEEN THE NATIVE AND DESIALYLATED FORMS OF HCG

Although the pregnancy hormone hCG has been extensively mapped immunoc hemically, few monoclonal antibodies have been produced to the unique COOH-terminal region of its beta-subunit (beta CTP). We now report the development and characterization of five such monoclonal antibodies. Three of these antibodies were developed to the synthetic peptide anal og of the hCG beta-(109-145) region coupled to diphtheria toroid, and two antibodies to a conjugate of bovine thyroglobulin and the peptide hCG beta-(115-145) prepared from hCG with its carbohydrate moieties in tact. The monoclonal antibodies raised against the synthetic peptide b ound hCG, desialylated hCG, and synthetic peptide to a similar extent, whereas antibodies generated to the natural hCG peptide did not bind to the synthetic peptide analog of the COOH-terminal peptide (beta CTP ) region or to desialyated hCG. These new monoclonal antibodies could distinguish between native and desialyated hCG in liquid phase immunoa ssays as well as by Western blots. They are highly specific reagents f or such Western blotting and were used for studies of a crude human pi tuitary gonadotropin preparation to demonstrate that it contained inta ct hCG beta without the internal peptide bond cleavages found in the s ubunit present in human blood and urine. Competition experiments using combinations of monoclonal antibodies and rabbit anti-beta CTP antise rum demonstrated that two epitopes exist within the beta-(115-145) reg ion of hCG, one of which depends on the presence of carbohydrate. In s ummary, the new monoclonal hCG beta CTP antibodies reported here can 1 ) discriminate between native and desialylated hCG, 2) identify hCG an d nicked hCG on Western blots, 3) provide an immunoaffinity purificati on tool for hCG, and 4) bind to two distinct epitopes on the beta CTP.