Recognition of HLA-A1 by murine monoclonal antibodies

Abstract: Balb/c mice were immunized with cells from the mouse mastocytoma line P815 transfected with an HLA-A1 gene. The splenocytes of the immunized mice were fused with cells from the murine myeloma NS-1. In an initial screening, supernatants of growing cultures were tested for their binding capacity to the immunizing P815/A1⁺ cells as well as to P815/A2⁺ cells. Three out of 756 hybrids produced antibodies which bound to P815/A1⁺ cells only. They were cloned and further analyzed for their binding reactivity to reference B-lymphoblastoid cells from the Tenth International Histocompatibility Workshop. One monoclonal antibody, designated 6B11, reacted only with HLA-A1⁺ cells, while the two other antibodies, 3G3 and 7F10, appeared to detect antigenic determinants shared by HLA-A1, A3, A11, A26, and A30 (3G3) and by HLA-A1, A3, A11, A26, A28 and A30 (7F10). Flow cytometric studies on B-lymphoblastoid cell lines as well as on a series of tumor cell lines, including melanoma and colon carcinoma lines, confirmed the specificity of these antibodies. Monoclonal antibodies 7F10 and 6B11 were found to be of the IgM class and 3G3 of the IgG1 class. By complement-dependent ⁵¹Cr release experiments it was further shown that the two IgM antibodies 7F10 and 6B11 were able to lyse all cell lines of the HLA-A1 haplotype tested.