Characterization of a major human antibody clonotype (1A) by monoclonal antibodies to combining site-associated idiotopes.
To identify major human antibody clonotypes with specificity to N-acetyl-D-glucosamine (GlcNAc), affinity-purified antibody preparations of different human individuals were analyzed by isoelectric focusing (IEF). A major clonotype (1A) was identified representing 7% of all anti-GlcNAc antibodies of one donor (no. 371). Anti-GlcNAc antibodies of donor 371 were used as immunogen to prepare monoclonal anti-idiotopic antibodies (mAb). Two anti-idiotopic mAb were specific for the major clonotype 1A, as shown by blotting of the anti-GlcNAc antibodies after IEF in thin-layer agarose to nitrocellulose filters and staining with either iodinated antigen or iodinated anti-idiotopic mAb. The two anti-idiotopic mAb 13F15 and 10F59 were further analyzed with respect to the antigenic determinant which they recognize. Both were directed to combining site-related determinants as shown by inhibition studies. Although 13F15 has a 500-fold higher binding capacity (relative affinity) equal amounts do not inhibit more than 50% of 10F59 binding, suggesting that the two mAb detect two closely related, but not identical, idiotopes in the antigen combining site of clone 1A. Although clonotype 1A is a unique antibody exclusively found in IEF of donor 371, the idiotope 1A.1, defined by mAb 13F15, is a recurrent determinant detectable on different anti-GlcNAc spectrotypes in various human sera.
D000117:Acetylglucosamine / Q000276:immunology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D000939:Epitopes / Q000032:analysis / Q000235:genetics / Q000276:immunology; D005944:Glucosamine / Q000031:analogs & derivatives*; D006801:Humans; D007130:Immunoglobulin Idiotypes / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D007525:Isoelectric Focusing; D051379:Mice; D051381:Rats
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007130:Immunoglobulin Idiotypes; D005944:Glucosamine; D000117:Acetylglucosamine