Analysis of the domain specificity of various murine anti-human IgM monoclonal antibodies differing in human B lymphocyte signaling activity.
The domain binding specificity of 19 murine anti-human IgM monoclonal antibodies (MoAbs) that have shown considerable heterogeneity in the transduction of stimulatory and inhibitory signals to B lymphocytes was evaluated by competition radioimmunoassays. Through the use of: (i) enzymatic fragments of IgM which each encompass more than a single CH domain, i.e. Fc5 mu and F(ab')2 mu, (ii) isolated single domains, C mu 2, C mu 3, and C mu 4, and (iii) mu heavy chain disease proteins, nine anti-IgM MoAbs were found to have C mu 1 domain specificity, five to have C mu 2 specificity, and five others to have C mu 4 specificity. Ineffective binding to isolated mu chain demonstrated that C mu 1-specific MoAbs were directed to epitopes which require light chain for expression. The lack of binding of the C mu 4-specific MoAbs to CNBr cleavage fragments of Fc5 mu suggest that the determinants recognized by these MoAbs may also be conformational in nature. Cross-inhibition analyses were used to determine the number of unique epitopes recognized by the anti-IgM MoAbs. Results from these experiments showed that: (i) eight of the nine MoAbs specific for C mu 1 likely bind to a single epitope, or very proximate epitopes, (ii) the five C mu 2-specific MoAbs recognize at least three distinct epitopes, and (iii) the five C mu 4-specific MoAbs each recognize a separate determinant. A comparison of the known B cell activating properties of these MoAbs with their specificity for the various segments of the IgM molecule indicate that mitogenicity cannot be attributed to selective binding to any one domain.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001402:B-Lymphocytes / Q000276:immunology*; D001667:Binding, Competitive; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000276:immunology; D007075:Immunoglobulin M / Q000276:immunology*; D051379:Mice
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000939:Epitopes; D007128:Immunoglobulin Fragments; D007147:Immunoglobulin Light Chains; D007075:Immunoglobulin M