Mixed leukocyte culture tests have been carried out on two hetero geneous but genetically defined backcross populations of rats to determine whether the reactions observed can provide the basis for histocompatibility matching. The experiments were designed so that only-one-way reactions could occur. Only when the donors of individual leukocyte mixtures differed at the important Ag-B histocompatibility locus was there any in vitro reactivity, and differences at this locus were invariably associated with the prompt rejection of skin homografts. Determination of compatibility at this locus proved to be important in that it facilitated the prolongation of survival of skin homografts by immunosuppressive therapy.