Production, purification and characterization of monoclonal antibodies specific to progesterone = 프로제스테론 단일클론 항체의 생산과 정제 및 특성연구

Abstract

Seven hybridoma cell lines, 051-01 (IgGi), 051-02 (IgGJ，051-03 (IgGj), 051-04 (IgM), 051-05 (IgG2a), 051-06 (IgM), and 051-07 (IgG汝) prodcing monoclonal antibodies against progesterone were established. Progesterone lla-hemisuccinate-bovine serum albumin conjugate was prepared using carbodiimide and then used as an immunogen. Spleen cells of Balb/c mice immunized with the immunogen were fused with mouse myeloma (P3-X63-Ag8. 653) cells. Hybridoma cells were screened by radioimmunoassay using 3H-progesterone. The clones 051-01，051-03, and 051-05 were purified using protein A-Sepharose 4B affinity chromatography utilizing their subtype characteristics. The purity of the isolated IgG was identified by SDS-PAGE. The progesterone binding capacity of the 5 monoclonal antibodies ranged from 7 to 72 pmole/mg IgG and the affinity constant ranged from 5.0X 107 M 1 to 4X108 M 1 by the Scatchard analysis method. Cross-reactivities of the monoclonal antibodies, 051-01 and 051-02 with other steroid hormones were compared. These antibody reactions were very specific to progesterone and sensitive in a competitive immunoassay.