Use of anti-idiotypic antibodies to establish that monoclonal antibody 7H11D6 binds to the alpha 2-macroglobulin receptor recognition site.
Studies were performed to determine if monoclonal antibody 7H11D6 binds to the region of alpha 2-macroglobulin (alpha 2M) that interacts with cell-surface receptors. F(ab')2 fragments prepared from this antibody delayed the in vivo clearance of alpha 2M-trypsin complexes from the murine circulation and blocked the in vitro binding of 125I-labeled alpha 2M-trypsin to rat kidney fibroblasts. Chemical modification studies revealed that the epitope region for 7H11D6 is sensitive to modification of the inhibitor with cis-dichlorodiammineplatinum (II). To determine if the epitope for 7H11D6 is comprised of residues involved in the alpha 2M receptor determinant, anti-idiotypic antibodies against 7H11D6 were prepared. A competitive enzyme-linked immunosorbent assay demonstrated that the anti-idiotypic IgG competed with alpha 2M-trypsin complexes for binding to 7H11D6. The anti-idiotypic IgG inhibited the binding of 125I-labeled alpha 2M-trypsin complexes to normal rat kidney fibroblasts, and a Kd of 194 pM for the binding of the anti-idiotype to these cells was derived from a fit of the data to a model involving a single class of sites. Binding of the anti-idiotypic IgG to the alpha 2M receptor provides unequivocal evidence that 7H11D6 is binding to residues within the receptor recognition site on alpha 2M and not merely to residues sufficiently close to that region to cause steric hindrance.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D001667:Binding, Competitive; D002478:Cells, Cultured; D002945:Cisplatin / Q000494:pharmacology; D000939:Epitopes / Q000032:analysis; D007130:Immunoglobulin Idiotypes; D007668:Kidney / Q000276:immunology; D007700:Kinetics; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D051381:Rats; D011971:Receptors, Immunologic / Q000187:drug effects / Q000276:immunology*; D000511:alpha-Macroglobulins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000939:Epitopes; D007130:Immunoglobulin Idiotypes; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D011971:Receptors, Immunologic; D000511:alpha-Macroglobulins; D002945:Cisplatin