In vitro immunization of human peripheral blood lymphocytes: establishmentof B cell lines secreting IgM specific for cholera toxin B subunit from lymphocytes stimulated with IL-2 and IL-4

In vitro immunization (IVI) techniques have a great potential in the produc tion of human monoclonal antibodies (MAbs) against various antigens. An IVI method of human peripheral blood lymphocytes (PBL) has been developed with a human lung adenocarcinoma cell line in our laboratory. Although several cancer specific human MAbs were successfully generated by using this IVI me thod, it was not available for soluble antigens, which prompted us to impro ve the method for generation of human MAbs against soluble antigens. IVI wi th soluble antigens was effectively caused by the addition of muramyl dipep tides, interleukin-2 and interleukin-4. It was found that the difference of sensitivity of lymphocytes depending upon donors could be overcome by find ing the optimal concentrations of IL-2 and IL-4. IVI of human PBL was perfo rmed with cholera toxin B subunit (CTB) and the immunized B cells were tran sformed by Epstein-Barr virus. Anti-CTB antibody was detected using an indi rect ELISA. B cells producing anti-CTB antibodies were directly cloned by a soft agar cloning method.