A new method for the development of human T-T cell hybrids without the use of HAT medium

We report here a new method for the development of human T-T cell hybrids by fusing mitogen- or alloantigen-stimulated T cells with non-mutagenized cells from human lymphoblastoid T cell lines. This method is based on a new selection procedure where the hybrids are separated from the parent T cell line on the basis of their ability to form colonies in soft agar. In contrast, cells from lymphoblastoid T cell lines Molt-4 and Jurkat do not form colonies in agar. Hybridoma colonies are retrieved from the agar plates, expanded in culture, screened by HLA-typing and appropriate functional tests and recloned several times by limiting dilution. HAT medium, which contains thymidine that appears to be toxic to the hybrids, is not used in our selection procedure. Using this method, we developed human T-T cell hybridomas (as determined by HLA-typing) producing B-cell growth factor (BCGF) either constitutively or after induction with Concanavalin A (Con A). Certain other T-T cell hybrids produced suppressor factor, significantly inhibiting proliferative responses of human peripheral blood mononuclear leukocytes to PHA, Con A and allogeneic cells in mixed lymphocyte culture.