Chelation challenge testing has been used to assess the body burden of various metals. The best-known example is EDTA challenge in lead-exposed individuals. This study assessed diagnostic chelation challenge with dimercaptosuccinic acid (DMSA) as a measure of mercury body burden among mercury-exposed workers. Former employees at a chloralkali plant, for whom detailed exposure histories were available (n = 119), and unexposed controls (n = 101) completed 24-hr urine collections before and after the administration of two doses of DMSA, 10 mg/kg. The urinary response to DMSA was measured as both the absolute change and the relative change in mercury excretion. The average 24-hr mercury excretion was 4.3 microg/24 hr before chelation, and 7.8 microg/24 hr after chelation. There was no association between past occupational mercury exposure and the urinary excretion of mercury either before or after DMSA administration. There was also no association between urinary mercury excretion and the number of dental amalgam surfaces, in contrast to recent published results. We believe the most likely reason that DMSA chelation challenge failed to reflect past mercury exposure was the elapsed time (several years) since the exposure had ended. These results provide normative values for urinary mercury excretion both before and after DMSA challenge, and suggest that DMSA chelation challenge is not useful as a biomarker of past mercury exposure.