Monoclonal antibodies against bovine adrenal cytochrome P-450(11 beta) and cytochrome P-450SCC. Their isolation, characterization and application to immunohistochemical analysis of adrenal cortex.
Seven monoclonal antibodies directed against bovine adrenocortical cytochrome P-45011 beta were isolated. They were found to be immunoglobulins each having distinct affinity to the antigen. Five clones had IgG1 heavy chains, whereas the other two had IgG2b heavy chains. All the clones had kappa light chains. All the monoclonal antibodies recognized a protein among mitochondrial proteins of bovine adrenal cortex, whose mol. wt, 50,000, was the same as that of cytochrome P-45011 beta. Among the monoclonal antibodies isolated, monoclonal antibody 258 recognized a protein of Mr 50,000 in rat adrenal mitochondria, a protein of Mr 47,000 in pig adrenocortical mitochondria, a protein of Mr 50,000 in guinea pig adrenal mitochondria, a protein of Mr 55,000 in rabbit adrenal mitochondria and a protein of Mr 50,000 in human adrenocortical mitochondria. These results suggest that each of these proteins, having a mol. wt around 50,000, recognized by monoclonal antibody 258 is very likely to be the cytochrome P-45011 beta of adrenocortical mitochondria of each animal species. Monoclonal antibody 42718 inhibited by 50% the steroid hydroxylation activity of cytochrome P-45011 beta. In addition to the monoclonal antibodies against cytochrome P-45011 beta, three monoclonal antibodies directed against bovine adrenocortical cytochrome P-450SCC were also isolated and characterized. Immunohistochemical staining of bovine adrenal cortex with the use of these two kinds of monoclonal antibodies revealed that the contents of both cytochrome P-45011 beta and cytochrome P-450SCC were greater in the zona fasciculata and zona reticularis than in the zona glomerulosa. Electron microscopical observation of immunoperoxidase-stained preparations confirmed the presence of these cytochromes in the mitochondria.
D000302:Adrenal Cortex / Q000201:enzymology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D002384:Catalysis; D002417:Cattle; D003101:Collodion; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006651:Histocytochemistry; D007120:Immunochemistry; D007074:Immunoglobulin G / Q000032:analysis
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D003101:Collodion; D003577:Cytochrome P-450 Enzyme System