We have shown that cytotoxic T cells can be polyclonally activated by a short exposure to complexes of polyadenylic:polyuridylic acid (poly A:U). Activation is optimal at a dose of 100 microgram/ml poly A:U and occurs during a 2 h incubation period in the absence of antigen. Killing of allogeneic, but not syngeneic, target cells can be observed after 12 h in culture and peaks after 21-24 h in the absence of any nonspecific binding ligand. The observed cytotoxicity is mediated by T lymphocytes and dose not require accessory macrophages or DNA synthesis for the activation or expression of effector functions. These results suggest that few requirements exist for the activation of cytotoxic T cells.