Inhibition by anti-Ia sera of guinea pig T lymphocyte proliferation induced by allogeneic macrophages (MLR) and NaIO4 or neuraminidase-galactose oxidase-treated macrophages has been investigated in order to identify the target cell upon which the antisera act. Anti-2 and anti-13 alloantisera were found to inhibit both MLR and aldehyde-induced T cell reactivity when directed against the specificity of the stimulatory macrophage. Little or no inhibition was observed when these antisera were directed against the T lymphocyte specificity when cultures were harvested at the time of peak proliferation. In addition, anti-2 serum was found to inhibit macrophage-lymphocyte rosette formation at 20 hr between neuraminidase-galactose oxidasetreated strain 2 macrophages and strain 13 lymphocytes. These findings demonstrate that inhibition of T cell proliferation can be produced by anti-Ia sera directed against the macrophage and raise the possibility that Ir gene products may function in part at the level of the macrophage.