Characterization of monoclonal antibodies recognizing alpha and beta subunits of human chorionic gonadotropin hormone

Human chorionic gonadotropin (hCG) hormone is required for maintenance of e arly pregnancy and is a potential marker in the diagnosis and prognosis of both pregnancy and trophoblastic diseases. Murine hybridomas were generated against purified hCG. Seven hybrid clones secreting antibodies against hCG molecule with IgG(1)/kappa subclass were established. The indirect ELISA r esult demonstrated that six MAbs (BEL-1 to BEL-6) recognized hCG in both ho le and free beta subunit form with negligible cross-reactivity against a cl osely related hormone, human luteinizing hormone (hLH). In this fusion, onl y one MAb (ALC-1) showed a cross-reaction with hLH, which designated an alp ha subunit specific. The outcome of Western blot ascertained that ALC-1 rec ognized the conformational epitope on alpha subunit of hCG at Mr 23 kDa ban d in nonreducing condition (NR). In contrast, epitopes belonging to ail MAb s recognized beta subunit of hCG were in linear peptide structure at Mr 34 kDa band (NR) and Mr 26 kDa band (R). These six MAbs were further character ized by using two beta subunit carboxy-terminal synthetic peptides (beta 10 9-119 and beta 109-145). Three of them (BEL-1, BEL-3, and BEL-4) recognized only epitope harboring in beta 109-145 fragment, the others recognized bot h types of the synthetic peptide. In order to select the most suitable MAbs specific to beta subunit of hCG for exploiting with ALC-1 in the sandwich- type immunometric assay, competitive ELISA was employed. Six individual MAb s specific to beta subunit of hCG were used to compete with biotinylated AL C-1 to evaluate the proximity of their epitopes on the hole form of hCG mol ecule. Of all six MAbs, BEL-5 depicted the lowest percent inhibition result , which indicated the bottom-most steric hindrance effect. Consequently, MA b BEL-5 will be the most appropriate antibody to utilize in concert with AL C-1 in place of devising a sandwich-type immunometric assay for measuring h olo-hCG level.