Cytotoxic T lymphocytes from healthy donors can be expanded to high numbers from the peripheral blood using combinations of anti-CD3 and anti-CD28 monoclonal antibodies (mAb). We investigated whether these antibodies could also be used to induce outgrowth of tumour-infiltrating lymphocytes (TIL) from tumour tissue. In the initiation phase of TIL culture immobilized anti-CD3 antibodies together with anti-CD28 mAb and low-dose interleukin-2 induced a rapid expansion of T cells from various human tumour tissues. The cultured cells showed high levels of cytotoxic T lymphocyte activity, but low levels of lymphokine-activated killer cell activity were generated. This study shows that TIL can be efficiently expanded from tumour tissue by combinations of anti-CD3 and anti-CD28 antibodies. This protocol for cell expansion in vitro may substantially reduce the time required to reach sufficient numbers of TIl for re-infusion to the patient.