Soft agar and limiting dilution techniques utilizing human T cell growth factor (TCGF) have been developed to clone human lymphoid cells with high cloning efficiency. TCGF has been used for the continued growth and expansion of lymphoid populations derived from single T cells, and the cytotoxic specificity of individual clones has been repeatedly examined. An analysis of clones derived from human lymphocytes sensitized to allogeneic determinants in vitro reveals significant polymorphism in lytic recognition when tested against HLA typed targets. This pattern of reactivity remained constant over the 2 months of continuous culture. These methods should be capable of dissecting and analyzing the fine specificity of the T cell response.