Monoclonal Antibodies to HLA–DRw Determinants

Abstract

Two monoclonal antibodies recognizing HLA–DRw (human Ia) antigens were produced. The DA2 antibody binds a monomorphic determinant, common to all specificities and Genox3.53 antibody binds to a cross-reacting site on the HLA–DRw1, 2 and 6 specificities. Both antibodies are IgGl and show complement dependent cytotoxicity only in the presence of rabbit anti-mouse IgG serum. Specificity of both antibodies for the HLA—DRw molecule was shown by inhibition of antibody binding by preincubation of antibody with detergent solubilized Ia from JY (HLA–DRw4, 6) cells and by preincubation of target cells with F(ab')₂ fragments of a rabbit anti-Ia serum. DA2 antibody reacted with all cells of human B cell origin tested and with peripheral blood lymphocytes of several primate species tested. Genox3.53 antibody bound only to human cells expressing HLA–DRw1, 2 or 6 antigens, giving a negative reaction with all primates tested. Genox3.53 antibody detected a split in the HLA–DRw6 specificity, showing reduced binding to the Daudi cell (HLA–DRw6) in comparison with binding to several other cell lines typed as HLA–DRw6, under saturating conditions. This low reactivity with Daudi was confirmed by absorption experiments. The ratio of DA2 binding to Genox3.53 binding to homozygous and heterozygous cell lines under saturation conditions was compared. Results suggested that, on some cell lines, DA2 might be reacting with a second population of human Ia antigens in addition to the HLA–DRw antigens. When a mixture of saturating concentrations of DA2 and Genox3.53 antibodies was tested for binding to cells under saturating conditions, the number of counts bound suggested the antibodies could bind simultaneously. Direct binding experiments showed that when each antibody was iodinated, its binding was not inhibited by preincubation with the other antibody, confirming that the DA2 and Genox3.53 determinants are distinct on the Ia molecule.