Affiliations: Department of Nephrology, Royal Melbourne Hospital, Parkville, Victoria, Australia
Note: [] Dr. Power's present address: Department of Medicine and Therapeutics, Polwarth Building, Aberdeen Royal Infirmary, Foresterhill, Aberdeen AB9 2ZD, Scotland
Note: [] Address requests for reprints to: A.J.F. d'Apice, M.D., Department of Clinical Immunology, St. Vincente Hospital, Fitzroy, Victoria 3050, Australia.
Abstract: We attempted to produce human monoclonal IgM antibodies to class II HLA antigens by in vitro immunization of human splenocytes with affinity purified HLA or irradiated whole lymphoblastoid B cells using the adjuvant muramyl dipeptide and IL-2. Two fusions where affinity purified HLA was used to stimulate the in vitro immunization produced no hybrids secreting antibodies against the stimulating antigen. However, 13.1% of 381 hybrids from eight fusions were positive by CELISA when whole cells were used to immunize. These antibodies were reactive with a wide range of cell types and were not directed to HLA. It proved difficult to characterize them, possibly because of low affinity. One reacted in Western blots with a 200 kD antigen, but this was exceptional. Most bound to a small percentage of cells in flow cytometry and did not lyse target cells in cytotoxicity assays. Control fusions were performed and similar antibodies were obtained from 2.6% of235 hybrids. These data suggest that in vitro immunization using the protocol outlined can increase the frequency of some antibodies, but that they may represent a very primitive lineage which does not have the specificity and affinity required to prove useful as diagnostic or therapeutic agents.
Keywords: human monoclonal antibodies, HLA, in vitro immunization
