Abstract
In vitro immunization and subsequent immortalization of peripheral blood cells of a multiparous woman has resulted in the production of a stable human mouse heterohybridoma, 5C2A2, secreting an HLA-A2/A28-specific human monoclonal antibody. Although possibly exposed to HLA-A2 by transfusions, the cell donor showed no HLA-A2-specific serum antibodies. The present protocol for in vitro immunization includes the elimination of suppressor cells from the responder cell population, the presence of irradiated allogeneic lymphocytes as a source of antigen, as well as stimuli--recombinant interleukin-2 and a B-cell specific nucleoside analogue--causing the proliferation of B lymphocytes, prior to immortalization. The ability of the antibody 5C2A2 to detect all known HLA-A2 subtypes, except A2.3, and A28, allows identification of the serological epitope on the HLA-A2 molecule. Application of this in vitro immunization method allows the production of a set of HLA monoclonal antibody-secreting human hybridomas, independent of the existence of serum HLA antibodies in the lymphocyte donors.
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