Combined therapy with a fixed-tumor cell vaccine and intratumoral injection of NK cells induced strong tumor regression of rat glioma. Rat 9L glioma cells were inoculated into syngeneic male rats at the flank (subcutaneous tumor model) or at the basal ganglia of the right hemisphere (intracranial tumor model). Rats were intradermally injected three times with vaccine comprising fixed 9L cells, IL-2- and GMCSF-microparticles, and tuberculin prior to (protective studies) or after (therapeutic studies) challenge with live 9L cells. In the protective studies, the vaccine alone achieved significant tumor growth inhibition and elongation of mean life span in both the subcutaneous and intracranial tumor models. No therapeutic effect was observed in the intracranial tumor model with the vaccine alone. However, intratumoral injection of rat NK cells strongly assisted the therapeutic effect of the vaccine in the brain tumor model and resulted in a statistically significant elongation of life span. We propose that intratumoral injection of NK cells may not only kill brain tumor cells directly, but also trigger a strong immune response in the focal lesion of the brain after vaccination.