Abstract A small proportion of human CD3+ T lymphocytes are known to co-express CD56, an antigen usually restricted in its expression to natural killer (NK) cells. Whereas the in vivo function of CD3+ CD56+ T cells remains unknown, we and others have previously shown that both in vitro and in vivo, these cells can mediate a significantly greater degree of MHC-unrestricted cytotoxicity against a variety of human tumor cells when compared to either CD3+ CD56− T cells or lymphokine activated killer (LAK) cells. While the mechanisms regulating the in vivo expansion of CD56+ T cells are not known, here we demonstrate the importance of CD2-mediated IL-12-dependent signals in the in vitro expansion of CD56+ T cells. Specifically, we show that activated monocytes provide a contact dependent factor (CD58/LFA-3) and a soluble factor (IL-12), both critical for the in vitro expansion of CD56+ T cells. The biological and therapeutic implications of these findings are discussed.