The consumption of trans-fatty acids adversely affects serum lipid levels. The relationship with the incidence of gallstone disease is unknown. This study prospectively examined consumption of trans-fatty acids in relation to the risk of gallstone disease in a cohort of 45 912 men. trans-Fatty acid consumption was assessed using a validated semiquantitative food frequency questionnaire. Newly diagnosed gallstone disease was ascertained biennially. During 14 years of follow-up, 2356 new cases of symptomatic gallstones were documented. After adjusting for age and other potential risk factors, compared with men in the lowest quintile of dietary intake of trans-fatty acids, the relative risk (RR) of gallstone disease for those in the highest quintile was 1.23 (95% confidence interval [CI], 1.04-1.44: P for trend, .03). Among individual trans-fatty acids, the RR for trans-oleic fatty acid, when extreme quintiles were compared, was 1.24 (95% CI, 1.06-1.45; P for trend, .02). Intakes of trans-palmitoleic fatty acid (RR, 1.09; 95% CI, 0.90-1.31), trans,trans 18:2 fatty acid (RR, 1.14; 95% CI, 0.96-1.34), and cis-trans 18:2 fatty acid (RR, 1.00; 95% CI, 0.86-1.16) were not significantly associated with the risk. The results suggest that a higher intake of trans-fatty acids modestly increases risk of gallstone disease.