Antiidiotypic antibodies raised against anti-prolactin (PRL) antibodies recognize the PRL receptor.
Antiidiotypic antibodies capable of recognizing the PRL receptor have been raised against antibodies to ovine PRL (oPRL) and rat PRL (rPRL). Anti-oPRL or anti-rPRL antibodies were induced in rats or rabbits, respectively, and the immunoglobulin G (IgG) fractions were isolated by affinity chromatography on a Protein A-Sepharose 4B column. Specific anti-oPRL antibodies were purified on an oPRL-Sepharose 4B affinity column. The specific antibodies (0.5 mg/rabbit) in Freund's complete adjuvant were injected into rabbits at 2- to 3-week intervals for 3 months. Antiidiotypic antibodies against rat anti-oPRL specifically inhibited [125I]iodo-oPRL binding to the immunizing anti-oPRL antibody. Membrane binding of the antiidiotypic antibodies was determined by [125I]iodo-Protein A precipitation. It was found to be significantly higher toward membrane preparations rich in PRL receptors, such as liver membranes from pregnant mice or from estradiol-treated male rats or prostate membranes from adult rats. This membrane binding by the antiidiotypic antibody was competitively inhibited by the immunizing anti-oPRL antibody, suggesting that the idiotypic antibody may share common determinants with the PRL receptor. Recognition of the PRL receptor by the antiidiotypic antibodies was also assayed by indirect binding studies. After preincubation of the antiidiotypic antibodies with the membranes, the resulting complex was precipitated and the amount of free antiidiotypic antibody remaining in the supernatant estimated according to its ability to inhibit [125I]iodo-oPRL binding to anti-oPRL IgG. The lowest degree of inhibition of [125I]iodo-oPRL binding was achieved after incubation of the anti-idiotypic IgG with liver membranes from pregnant mice, while the inhibitory capacity was about 5-fold higher subsequent to parallel incubations with the same membranes, which had previously been heated for 30 min at 65 C, or with membranes of male rat liver, demonstrating a direct correlation between the binding of the antiidiotypic antibody to the membranes and the PRL receptor content of the membranes. Furthermore, significant and dose-dependent inhibition of [125I]iodo-oPRL binding to its receptors in various PRL receptor-rich membrane preparations from rats and rabbits was demonstrated with antiserum of rabbits immunized with rabbit anti-rPRL IgG. These results suggest that effective and specific anti-PRL receptor antibodies can be produced using the antiidiotypic antibody procedure, thus avoiding the necessity of isolating and purifying the PRL receptor itself.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology*; D000918:Antibody Specificity; D002462:Cell Membrane / Q000276:immunology; D005260:Female; D007106:Immune Sera / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D007130:Immunoglobulin Idiotypes / Q000276:immunology*; D008099:Liver / Q000276:immunology; D008297:Male; D051379:Mice; D011247:Pregnancy; D011388:Prolactin / Q000276:immunology*; D011467:Prostate / Q000276:immunology; D011817:Rabbits; D051381:Rats; D011919:Rats, Inbred Strains; D011956:Receptors, Cell Surface / Q000276:immunology*; D011981:Receptors, Prolactin
D000888:Antibodies, Anti-Idiotypic; D007106:Immune Sera; D007074:Immunoglobulin G; D007130:Immunoglobulin Idiotypes; D011956:Receptors, Cell Surface; D011981:Receptors, Prolactin; D011388:Prolactin