OBJECTIVE: Sex differences in pain are well established, with women reporting greater incidence of clinical pain and heightened responsivity to experimental pain stimuli relative to men. Sex hormones (ie, estrogens, progestins, androgens) could contribute to extant differences in pain sensitivity between men and women. Despite this, there has been limited experimental research assessing the relationship between pain and sex hormones. The purpose of this study was to extend previous research and examine the association between sex hormones and nociceptive processing in healthy women. MATERIALS AND METHODS: A total of 40 healthy women were tested during the mid-follicular, ovulatory, and late-luteal phases of the menstrual cycle (testing order counterbalanced). Salivary estradiol, progesterone, and testosterone were collected at each testing session and pain was examined from electrocutaneous threshold/tolerance, ischemia threshold/tolerance, and McGill Pain Questionnaire-Short Form ratings of noxious stimuli. Nociceptive flexion reflex threshold was assessed as a measure of spinal nociception. RESULTS: Overall, there were no significant menstrual phase-related differences in pain outcomes. Nonetheless, variability in testosterone (and to a lesser degree estradiol) was associated with pain; testosterone was antinociceptive, whereas estradiol was pronociceptive. No hormone was associated with nociceptive flexion reflex threshold. DISCUSSION: Although future research is needed to replicate and extend these findings to clinical populations (ie, chronic pain, premenstrual dysphoric disorder), results from the present study indicate that menstrual phase-related changes in sex hormones have minimal influence on experimental pain. However, individual differences in testosterone may play a protective role against pain in healthy women.