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.