Exposure to environmental tobacco smoke (ETS) has been associated with adverse health consequences. In this study, the association between self-reported ETS exposure and serum cotinine levels was examined. As part of the population-based Epidemiology of Hearing Loss Study in Beaver Dam, WI, self-reported data on ETS were obtained at the 5-year follow-up examination (1998-2000; n = 2800). Serum cotinine levels were obtained on 643 of these participants (53-75 years old). Serum cotinine levels increased with reported number of hours of recent ETS exposure. Most (95.3%) participants who reported being smokers had serum cotinine levels >15 ng/mL, compared with only 2.7% of nonsmokers. Usual ETS exposure among nonsmokers was assessed with a brief questionnaire quantifying exposure at work, at home, and in social settings, and then classified into three levels of exposure. Those reporting little or no exposure had a geometric mean cotinine level of 0.06 ng/mL (95% confidence interval (CI): 0.05, 0.07), compared with 0.14 (95% CI: 0.09, 0.22) and 0.63 (95% CI: 0.36, 1.10) for those reporting moderate and high ETS exposure, respectively (F-test for trend: P < 0.001). These results suggest that the questionnaire can be used to distinguish relative levels of exposure to ETS.