Tumor infiltrating lymphocytes (TIL) were isolated by centrifugal elutriation from C4 mouse mammary tumors and characterized with regard to phenotype and natural killer (NK) activity. Tumors that had arisen spontaneously in preneoplastic hyperplastic alveolar nodules and tumors that had been passaged one to two times in either naive or presensitized mice were studied. Mice were sensitized by limited s.c. tumor growth and subsequent surgical removal of the tumor. The total numbers of T or B cells in the infiltrates were similar in spontaneous tumors and in passaged tumors from either naive or sensitized mice. The ratio of L3T4-positive to lyt-2-positive cells was reduced, however, from 1.10 +/- 0.2 in spontaneous tumors to 0.53 +/- 0.28 or 0.48 +/- 0.04 in passaged tumors from untreated or sensitized mice. The site of tumor implantation, whether intramammary fat pad or s.c., did not affect the profiles of the infiltrates. The TIL from both spontaneous and passaged tumors demonstrated enhanced NK activity relative to peripheral lymphoid cells. The TIL of passaged tumors sensitized mice, however, had lower NK activity than those from naive mice.