Human monoclonal antibody to purified protein derivative of tuberculin produced by hybrids constructed with Epstein‐Barr virus‐transformed B lymphocytes and mouse myeloma cells

Abstract

A method of producing human monoclonal antibody by combining somatic cell hybridization technology with the capability of Epstein-Barr virus (EBV) to transform human B lymphocytes is described. Peripheral blood lymphocytes from a donor with positive tuberculin skin test reaction were transformed by EBV and then tested for antibody production to mycobacterial purified protein derivative (PPD) by an enzyme-linked immunosorbent assay. Two EBV-transformed lymphoblastoid cell lines making IgM antibodies to PPD were obtained. One of these cell lines was fused by polyethylene glycol with a murine hypoxanthine-guanine phosphoribosyl transferase-deficient myeloma cell line that had been selected for resistance to ouabain. The human-mouse hybrids were selected in ouabain-containing HAT medium and 11 heterohybridomas producing IgM antibody to PPD were obtained. one of these was cloned by limiting dilution with efficiency at least 20-fold higher than parent EBV-transformed cell line. Heterohybridoma subclones reached levels of IgM antibody as high as 75.0 μg/ml of culture medium, whereas IgM production of EBV-transformed B cell clones ranged between 3.0 and 4.0 μg/ml.