FACTORS AFFECTING PRODUCTION OF ANTIBODIES TO PROSTATE ANTIGENS BY IN-VITRO-PRIMED HUMAN SPLENOCYTES

Efficiency of immunization of splenocytes in vitro by three kinds of p rostate antigen has been studied. intact cells of the human prostate c ell line LNCaP, a membrane preparation (PMA) of LNCaP cells, and prost ate specific antigen (PSA) were used as antigens. Three different sche mes of in vitro immunization were tested: male spleen cells vs female; single donor spleen cells vs. mixed lymphocyte culture (MLC); additio n of exogenous IL-2 vs. no lymphokine addition. The supernatant antibo dies were tested for reactivity to the immunizing antigen by ELISA. Su bsequently hybridomas were generated by fusing the primed lymphocytes to a heteromyeloma cell line, K6H6/B5. Only antigen specific IgM respo nses could be detected. Intact LNCaP cells induced the highest respons es from mixed lymphocyte cultures. PMA also induced the highest respon ses from mixed lymphocyte cultures of male origin, whereas both single donor and mixed donor spleen cell cultures of female origin responded to PMA. However, anti-PMA responses by mixed lymphocyte cultures of f emale cells were significantly augmented by addition of IL-2. PSA only induced specific responses from mixed female splenocyte cultures supp orted with IL-2. Several anti-PMA and -PSA antibodies were generated a fter somatic fusion of the in vitro primed cells. Two monoclonal IgG a ntibodies from LNCaP primed spleen cells could be competitively inhibi ted with tumor membrane antigen.