A recent subtractive cDNA cloning study in rats demonstrated an unexpected increase in expression of the proteinase inhibitor, cystatin C in the spinal cord during acute peripheral inflammation, suggesting this protein may be involved in the pathogenesis of persistent pain. A subsequent study of 10 women suggested that prolonged labor pain resulted in increased cystatin C concentrations in cerebrospinal fluid, and that this could be used as a biomarker for pain. To confirm and extend these observations, we measured cystatin C concentrations in cerebrospinal fluid in 131 subjects: 30 normal volunteers without pain, 25 women at elective cesarean section without pain, 60 women in labor with severe pain, and 16 patients with chronic neuropathic pain and tactile allodynia. The median cystatin C concentration in normal volunteers, 2.2 microg/ml, was similar to that previously reported by multiple investigators, and cystatin C concentrations were increased in women in labor (3.9 microg/ml). However, contrary to the previous report, cystatin C concentrations in laboring women with pain did not differ from those of pregnant women without pain (3.7 microg/ml). There was no relationship between duration of painful labor and cystatin C concentration. Patients with neuropathic pain had similar cystatin C concentrations (2.4 microg/ml) to controls. Logistic regression analysis indicated that cystatin C concentrations could not be used to reliably predict the presence of pain in either acute or chronic settings. These data suggest that cystatin C concentration in cerebrospinal fluid is an unreliable diagnostic marker for pain in humans.