A new cloning method for antibody-forming lymphoblastoid cells. Increase in cloning efficiency by inclusion of human fibroblasts into semisolid agarose growth layer.

We report the development of a method for cloning human EBV-transformed cells which has greater efficiency than techniques used presently. In this new method lymphoblastoid cells are cultured in semisolid agarose in close physical association with human fibroblasts. The results indicate a 10-fold increase in the cloning efficiencies. The average cloning efficiency, depending on the age of cell lines, was from 1 to 14%, and colonies appeared 7-9 days sooner than in the traditional soft agarose method. The new method has allowed us to develop several stable lymphoblastoid cell lines producing antibody cytotoxic to human B lymphocytes. This method may make it more practical to obtain monoclonal human antibodies from lymphoblastoid cell lines which had previously been unstable due to heterogeneity.