title: Production of human monoclonal antibodies to B lymphocyte cell surface antigens by in vitro immunization and human-human hybridoma technology.
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% of 235 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.
authors.affiliation: Department of Nephrology, Royal Melbourne Hospital, Parkville, Victoria, Australia.
authors.forename: D A
authors.initials: DA
authors.lastname: Power
authors.forename: V
authors.initials: V
authors.lastname: Gerkis
authors.forename: A J
authors.initials: AJ
authors.lastname: d'Apice
mesh_terms: D000119:Acetylmuramyl-Alanyl-Isoglutamine; D000276:Adjuvants, Immunologic; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D000915:Antibody Affinity; D000918:Antibody Specificity; D001402:B-Lymphocytes / Q000276:immunology*; D002459:Cell Fusion; D002460:Cell Line; D006684:HLA-DR Antigens / Q000276:immunology*; D006801:Humans; D006825:Hybridomas / Q000276:immunology*; D007114:Immunization / Q000379:methods*; D007075:Immunoglobulin M / Q000096:biosynthesis*; D007376:Interleukin-2 / Q000494:pharmacology; D007518:Isoantibodies / Q000096:biosynthesis* / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D000911:Antibodies, Monoclonal; D006684:HLA-DR Antigens; D007075:Immunoglobulin M; D007376:Interleukin-2; D007518:Isoantibodies; D000119:Acetylmuramyl-Alanyl-Isoglutamine