A human monoclonal antibody that detects HLA‐A1, A23 and A24 antigens

Abstract

We report the production and characterization of a human monoclonal IgM (mu, kappa) antibody recognizing the HLA A1, A23 and A24 antigens. B lymphocytes obtained from a multiparous Japanese woman were transformed in vitro by Epstein-Barr virus, screened with an immune adherence assay, and fused with a murine myeloma cell line, P3-X63-Ag8.653. After subcloning by limiting dilution three times, a stable antibody-secreting hybridoma cell line, 4-35-7, was identified. The culture supernant had a titer of 1:32-64 against each of A1-, A23- and A24-positive lymphocyte panels, and showed complete correlation (r = 1.00) with the A1, A23 and A24 antigens on a lymphocyte panel of 287 unrelated, class I HLA-typed donors by the NIH cytotoxicity assay. Monoclonality of the antibody was ensured by Southern blot analysis of the human immunoglobulin heavy chain gene of 4-35-7. In view of the published data on HLA class I nucleotide sequences, the antibody may recognize an antigeneic determinant including two amino acid residues, Asp-166 and Gly-167, in the alpha 2 helix of the class I molecule that are specific for A1, A23 and A24 so far analyzed.

Generation and characterization of a human monoclonal IgG antibody specific for HLA‐B12

“…As a source for this purpose, alloantisera from multiparous woman, mouse monoclonal antibodies (mAb) and human mAb are used. In the last few years, a series of specific human anti-HLA class I mAb was published (9)(10)(11)(12)(13)(14)(15)(16). The uniformly used protocol for producing these mAb was Epstein-Barr-Virus (EBV) transformation of peripheral B lymphocytes and fusing of anti-HLA class I antibody-producing lymphoblastoid cell lines (LCLs) with myeloma cells (17).…”

Nucleotide sequences of the variable regions of a human monoclonal antibody against HLA‐A1, A23, and A24

“…recognized an antigenic determinant including two amino acid residues, Asp-166 and Gly-167 in the a? helix of the class I molecule, which were specific for A 1, A23 and A24 (8).…”