The Generation of Hybridomas Secreting Human Monoclonal Antibodies Reactive with Type II Collagen

Abstract

Human monoclonal antibodies reactive with type II collagen were obtained from patients with rheumatoid arthritis and relapsing polychondritis by fusion of cells with the human-mouse myeloma analogue HMMA2.11 TG/O. Direct fusion of peripheral blood or bone marrow mononuclear cells was unsuccessful in obtaining antibody producing hybridomas although fusion effiency was high (1 hybridoma per 25,400 mononuclear cells fused). Polyclonal, Epstein Barr Virus transformed B cell lines derived from peripheral blood and bone marrow mononuclear cells after direct transformation, in vitro secondary immunization, and/or CD8 depletion were found to produce antibodies that reacted with type II collagen. Antibody producing EBV transformed B cells were fused with the human-mouse myeloma analogue HMMA2.11 TG/0 and six separate, stable IgM antibody producing hybridomas obtained. These results demonstrate the difficulty in obtaining human monoclonal autoantibodies, particularly of IgG isotypes, using readily accessible sources of cells in patients with chronic autoimmune diseases without expansion and preselection.