A new set of monoclonal antibodies to human MHC class II alpha chains demonstrates that most alpha epitopes are inaccessible on the living cell surface.
When mice were immunized with a mixture of human MHC class II alpha and beta glycoprotein chains, the predominant antibody response was anti-alpha, and from a subsequent fusion experiment over 60 hybridomas showing anti-alpha activity were generated, compared with 11 anti-beta secretors. These findings contrast with the relative paucity of anti-alpha monoclonals described previously. Use of a miniaturized Western blot screening protocol was a critical factor in the present study since the anti-alpha monoclonals do not bind to the surface of living B cells and would therefore be missed in conventional screening assays. After glutaraldehyde fixation of target B lymphocytes or B-cell lines, the majority of anti-alpha monoclonals do react in a radio-immunobinding assay, although none binds as strongly as pan-reactive anti-beta chain antibodies. This suggests that the immunogenic epitopes of alpha chains are normally concealed by the three-dimensional folding of the alpha beta dimer. The anti-alpha monoclonals were all monomorphic but varied in the extent of their reactivity with alpha chains separated on one-dimensional and two-dimensional IEF gels. The most reactive antibodies identified up to seven distinct components among mature class II antigens from solubilized cell membranes.
D000818:Animals; D000911:Antibodies, Monoclonal; D000954:Antigens, Surface / Q000032:analysis; D001402:B-Lymphocytes / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D006681:HLA-D Antigens / Q000032:analysis*; D006801:Humans; D007525:Isoelectric Focusing; D051379:Mice; D008807:Mice, Inbred BALB C
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D000939:Epitopes; D006681:HLA-D Antigens