Isolation of steroid receptor binding protein from chicken oviduct and production of monoclonal antibodies.
Previous studies have shown that the molybdate-stabilized progesterone receptor from the chick oviduct contains a nonhormone binding component with a molecular weight of 90 000. This protein has also been shown to be associated with some other molybdate-stabilized steroid receptors of the oviduct. In order to access this larger pool of the receptor binding protein, we have developed an isolation procedure based on the observation that the protein is selectively shed from proteins adsorbed to heparin-agarose when molybdate is removed. The protein obtained by this procedure is shown to be the same as that isolated from affinity-purified progesterone receptor as compared by protease digestion and one-dimensional peptide mapping. Four immunoglobulin G secreting hybridoma cell lines were generated against the 90 000-dalton antigen. All of the antibodies recognize the 90 000-dalton protein obtained by electrophoretic transfer from sodium dodecyl sulfate-polyacrylamide gels. In addition, two of the antibodies complex the molybdate-stabilized progesterone receptor as demonstrated by sedimentation analysis on sucrose gradients. One of these antibodies was used to show the presence of the 90 000-dalton component in molybdate-stabilized glucocorticoid and androgen receptors and also to show its presence in brain, liver, and skeletal muscle, but not in serum.
D000818:Animals; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins / Q000276:immunology / Q000302:isolation & purification*; D002645:Chickens; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006825:Hybridomas / Q000276:immunology; D051379:Mice; D008970:Molecular Weight; D010057:Oviducts / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis; D011980:Receptors, Progesterone / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D010446:Peptide Fragments; D011980:Receptors, Progesterone