We assayed thymidylate synthase (TS) activity in normal and malignant gynecologic tissues. TS activity in the normal cervix, secretory endometrium, and ovaries from a total of 61 patients was 1.00 +/- 0.52 pmol/g tissue (mean +/- SD). Thus the upper limit (mean + 2SD) of TS activity in normal gynecologic tissue was 2.00 pmol/g tissue. TS activity was generally higher in malignant tissue than in normal tissue, and particularly high activity was detected in cervical cancer. TS activity in endometrial cancer was relatively low, but activity in poorly and moderately differentiated tumors was significantly elevated compared to well differentiated tumors. These findings suggest that cervical cancer is a highly proliferative tumor and that in endometrial cancer, cell proliferation is more rapid in poorly differentiated tumors than in well differentiated ones.