We describe the generation and immunotherapeutic properties of syngeneic immune T cells activated in situ against the lymphoma variant ESb which metastasizes to multiple organs (liver, lung, spleen, bone) and can develop sophisticated immune escape mechanisms. Peritoneal effector cells (PEC) generated from intra-pinna tumor immunized and i.p. restimulated mice were able to transfer protective immunity in the absence of exogenous cytokines into normal or 5Gy irradiated or SCID mice which had been injected either s.c. or i.v. with 5x10(3) to 10(5) metastatic ESb tumor cells. Protective immunity was specific for ESb cells and could be followed by growth retardation of s.c. tumors as well as by prevention or retardation of death from metastases. Expression of protective immunity was independent of host T cells since it was similarly expressed in SCID-mice and sublethally irradiated mice. Preincubation in vitro for 24 h of the PEC before adoptive transfer led to pronounced decrease of protective immunity while the tumor specific cytotoxic T lymphocyte (CTL) activity was maintained or even enhanced. These results demonstrate i) that protective immunity in vivo requires more than tumor specific cytotoxic activity and ii) that in vitro culture of immune cells can change their in vivo functional properties.