Generation of monoclonal antibodies to human chorionic gonadotropin by a facile cloning procedure.

Abstract

A facile hybridoma procedure has been used to generate monoclonal antibodies to alpha- and beta-subunits of human chorionic gonadotropin (HCG). The procedure is based on the method of Davis et al. (1982, J. Immunol. Methods 50, 161) and involves the use of a semisolid medium containing methylcellulose for the initial cloning of hybrid cells following immunization and cell fusion. Seven to ten days after cell fusion, viable hybrid clones were removed for subculture in a liquid medium containing RPMI 1640 and 15% fetal calf serum. Initial screening of hybrid cell lines that secrete antibodies to HCG was performed on microplate enzyme-linked immunoassay (ELISA) using HCG-coated microtiter plates. The specificity of these antibodies to either alpha- or beta-subunits was determined by the sodium dodecyl sulfate-gel/protein blot radioimmunobinding method which separates alpha- and beta-subunits of HCG on nitrocellulose strips for radioimmunobinding assay. As a result of this study, it has been possible to generate about 272 hybrid cell lines that secrete antibodies reacting with either the alpha- or beta-subunit of HCG in about 5 weeks. The association constants and cross-reactivities to luteinizing hormone for some of the HCG monoclonal antibodies were determined. The high affinity and specificity of these monoclonal antibodies permit their clinical application in a sensitive sandwich solid-phase enzyme-linked and radioimmunoassay of HCG.