Characterization of high affinity monoclonal antibodies against the luteinizing hormone-releasing hormone

Four hybridoma clones (BKL1, BKL2, BKL5 and BKL6), secreting monoclonal antibodies against the decapeptide luteinizing hormone releasing hormone (LHRH), were obtained by the fusion of Sp 2/0-Ag14 myeloma cells with spleen cells of a Balb/k mouse immunized with a conjugate of thyroglobulin-LHRH. All four monoclonal antibodies belong to the IgG1 subclass. The antibodies cross-reacted in RIA from 9.3 to 21% with 4-10 LHRH and less than 1% with 7-10 LHRH, 1-3 LHRH and LHRH-OH; 4-6 LHRH showed no cross-reaction. A RIA based on the BKL2 antibody and able to detect 4 pg LHRH per tube was developed. An extract of rat hypothalamus was submitted to Sephadex G50 separation and a single peak of LHRH immunoreactivity corresponding to synthetic LHRH, was detected with the antibody BKL2. When this material was further analyzed by HPLC, we found that the major peak of immunoreactivity co-migrated with synthetic LHRH. The data shows that the antibodies should be useful tools for biochemical and physiological studies on LHRH.