Comparison between different rabbit antisera against the glucocorticoid receptor.
Seven antisera against the glucocorticoid receptor (GR), raised in different rabbits immunized with highly purified (in case of five rabbits apparently homogeneous) preparation of GR from rat liver cytosol, were compared concerning titer and cross-reactivity. The titers of protein A-purified antisera (10 mg/ml) were in the range 1:100-1:320 as measured by enzyme-linked immunosorbent assay, ELISA, (defined as the dilution giving 50% of maximum absorbance). All seven antisera bound to the rat GR with a Stokes radius of 6.1 nm, but no antiserum reacted with the proteolytically induced steroid binding domain with Stokes radius 3 nm. However, the antigenic determinant(s) of the non-ligand-binding domain(s), split off from the steroid binding domain, is preserved following digestion with alpha-chymotrypsin or trypsin, respectively, since immunoactivity is still detectable by ELISA. Only two of four antisera tested cross-reacted with the GR from human lymphocytes. The same two antisera cross-reacted with chick embryo liver GR. Four out of four antisera tested cross-reacted with mouse liver GR as well as with rabbit lung GR. For these antisera, antibody binding to the GR prior to steroid- or DNA-binding did not influence the ability of the GR to interact with the ligand or DNA-cellulose, respectively. No difference regarding avidity of the antisera for activated or non-activated GR was observed. Furthermore, none of the antisera tested cross-reacted with the estrogen, progestin, androgen or mineralocorticoid receptors in rat. These findings indicate that the antisera from different rabbits raised against the same antigen all react with a certain domain of the rat GR, but show species differences as well as receptor class specificity.
D000818:Animals; D000936:Antigen-Antibody Complex; D003600:Cytosol / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D007106:Immune Sera; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000276:immunology / Q000378:metabolism*; D011987:Receptors, Steroid / Q000378:metabolism*; D014222:Triamcinolone Acetonide / Q000378:metabolism
D000936:Antigen-Antibody Complex; D007106:Immune Sera; D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D014222:Triamcinolone Acetonide