Optimizing production of human monoclonal IgG antibodies by in vitro-primed human PBMC: influence of CD56+ NK cell depletion.
Freshly isolated human peripheral blood mononuclear cells (PBMC) were immunomagnetically depleted of CD56+ cells. When these CD56- PBMC populations were cultured in the presence of autologous donor serum, polyclonal activation with IL-2 and pokeweed mitogen (PWM) generally resulted in exclusive production of IgG antibodies. Fusion with SP2/O-Ag14 mouse myeloma cells was highly efficient and yielded a great number of IgG-producing heterohybridomas. These conditions were used for in vitro immunization with viable human HT29 tumor cells. After fusion, an increase in hybridoma clones producing IgG monoclonal antibodies (MAb) with HT29 specificity showing a higher portion of MAb binding to the surface of viable HT29 cells was recorded. This immunizing efficiency was not observed with HT29 membrane protein fractions or HT29 proteins integrated into ISCOM particles. Investigations with human anti-alpha Gal antibodies showed that the IgG antibodies produced by the human/mouse heterohybridomas did not contain the mouse-specific Gal alpha 1-3Gal epitope.
Friedrich-Miescher-Laboratorium, Max-Planck-Gesellschaft, Tübingen, Germany.
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Puhlmann
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Anderer
D000276:Adjuvants, Immunologic / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000378:metabolism; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D019002:CD56 Antigen / Q000502:physiology*; D015179:Colorectal Neoplasms; D006031:Glycosylation; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007074:Immunoglobulin G / Q000096:biosynthesis*; D018189:Immunomagnetic Separation; D007694:Killer Cells, Natural / Q000276:immunology*; D007963:Leukocytes, Mononuclear / Q000276:immunology* / Q000378:metabolism; D008212:Lymphocyte Depletion; D051379:Mice; D014407:Tumor Cells, Cultured
D000276:Adjuvants, Immunologic; D000911:Antibodies, Monoclonal; D019002:CD56 Antigen; D007074:Immunoglobulin G