We have reported earlier that pretreatment of mice with neem leaf preparation (NLP) causes prophylactic growth inhibition of murine Ehrlich's carcinoma (EC) and B16 melanoma. Using adoptive cell transfer technology, here we have established that NLP-mediated activation of immune cells may be involved in tumor growth restriction. Mononuclear cells from blood and spleen of NLP-activated Swiss and C57BL/6 mice causes enhanced cytotoxicity to murine EC cells in vitro. Fractionation of spleen cells exhibited greater percentage of tumor cell lysis in macrophage and B-cell-depleted NK and T-cell-rich fractions. Flow cytometric analysis revealed in both blood and spleen, NK cells (DX5+ or NK1.1+) and NK-T cells (CD3+/DX5+ or CD3+/NK1.1+) were increased in number in Swiss, C57BL/6 and athymic nude mice after pretreatment with NLP. NLP-stimulated spleen cells showed greater secretion of TNFalpha and IFNgamma. Thus, NLP-activated NK and NK-T cells in mice may regulate tumor cell cytotoxicity by enhancing the secretion of different cytotoxic cytokines.