The T-lymphocyte differentiation alloantigens, A.R.T.-1 and A.R.T.-2 were used to follow the functional development of these cells in the rat. Delayed hypersensitivity and lymphocytes cytotoxic for allogeneic tumor cells were present in adoptively transferred recipients by 3 wk postirradiation and fetal liver reconstitution. However, these animals did not regain appreciable GvH potential until 7.5 wk postreconstitution. Normal percentages of A.R.T-1 lymphocytes were observed at all times in the spleen and lymph nodes of these animals, whereas, below normal numbers of A.R.T-2 lymphocytes were observed up to 7.5 wk postreconstitution. Depletion experiments using specific anti-A.R.T. sera demonstrated that the proliferative response to PHA and the cytotoxic lymphocyte response to alloantigens were reduced after treatment with anti-A.R.T.-1, but not with anti-A.R.T.-2. In contrast, the GvH potential of lymph node cells was reduced by treatment with either anti-A.R.T.-1 or anti-A.R.T.-2 serum plus complement. The use of these A.R.T. alloantisera in the rat has greatly facilitated the establishment of differentiation patterns for functional T lymphocytes and has resulted in the association of specific functional T-cell subpopulations with characteristic surface alloantigens.