Production of monoclonal antibodies against human growth hormone releasing hormone and their use in an enzyme-linked immunosorbent assay (ELISA)

Abstract

Two murine monoclonal antibodies (mAbs) specific for human growth hormone releasing hormone (GHRH-44-NH2) were produced from a fusion of spleen cells from a BALB/c mouse immunized with GHRH-conjugated BSA with SP 2/0 myeloma cells. The antibodies were of the IgG1 kappa, and IgG2b-kappa isotypes. The binding of both antibodies to GHRH-coated plates was inhibited by a 30-44 amino acid fragment but not by a 1-26 fragment. Thus, both antibodies are directed against the carboxy terminus of the peptide. Furthermore, both antibodies bind to the same epitope on the 30-44 amino acid portion since they cross-inhibit each other's binding to intact GHRH. Using these mAbs, a direct binding GHRH enzyme-linked immunosorbent assay (ELISA) was developed which had a least detectable dose of 30 pg. The availability of these antibodies and their use in ELISA methodology permits consistent and specific detection of GHRH in a non-isotope assay. They should prove of value in screening acromegalic patients for ectopic sources of GHRH secretion and in studies of ontogenic analysis of GHRH production.