We have prepared anti-Ly-6.2 by immunization of (CBA/Ca X A-Thy-1a)F1 with AKR/Crc lymphocytes and used the antiserum to study heterogencity of peripheral T lymphocytes. The Ly-6 alloantigen is found on all activated cytotoxic T cells studied, directed to a variety of different target antigens. The antiserum reveals heterogeneity among the precursors of T killer cells, i.e. allogeneic precursors are Ly-6-, whereas the generation of cytotoxic cells to hapten-modified syngeneic cells and xenogeneic cells is partially abrogated by serum pretreatment. Precursors of some in vivo primed responses are also Ly-6+. These findings indicate that the expression of Ly-6 antigen is, at least, partially related to the stage of T cell activation. The in vitro proliferative response of T cells to allogeneic Ia antigens is substantially reduced by pretreatment with anti-Ly-6.2 serum suggesting that this antiserum may be a useful tool for the study of heterogeneity in the Ly-1+ pool of T cells.