Results of a randomized controlled trial have suggested a protective effect of selenium against prostate cancer. Few other prospective studies have been conducted to confirm or refute this. The association between prostate cancer and baseline toenail selenium level was evaluated in the Netherlands Cohort Study, conducted among 58,279 men, aged 55-69 years at entry. In September 1986, the cohort members completed a questionnaire on risk factors for cancer and provided toenail clippings for determination of baseline selenium status. After 6.3 years of follow-up, 540 incident prostate carcinoma cases and 1,211 subcohort members with complete toenail selenium data were available for case-cohort analyses. In multivariate survival analysis, an inverse association between toenail selenium level and prostate cancer risk was observed. Incidence rate ratios in increasing selenium quintiles were 1.00 (ref), 1.05, 0.69, 0.75, and 0.69 (95% confidence interval, 0.48-0.99), respectively (P-trend = 0.008). This association persisted after exclusion of cases diagnosed during early follow-up. The inverse association was more pronounced in ex-smokers than current smokers, and unclear in never-smokers. Analysis of effect modification by intake of antioxidant vitamins C, E, and the carotenoids α-carotene, β-carotene, β-cryptoxanthin, lycopene, and lutein/zeaxanthin showed a strong, significant interaction with β-cryptoxanthin, and to a lesser extent with vitamin C. These results confirm the hypothesis that higher selenium intake may reduce prostate cancer risk. Future research on optimum dose level is needed.