Background: Although trans fatty acids (TFAs) may increase the risk of dyslipidemia and coronary artery disease (CAD), limited data are available on their association with heart failure (HF). Objective: Our goal was to assess associations of plasma and dietary TFAs with HF and CAD. Design: We used a prospective, nested case-control design to select 788 incident HF cases and 788 matched controls from the Physicians' Health Study for biomarker analyses and a prospective cohort for the dietary analyses. Plasma fatty acids were assessed by using gas chromatography, and dietary intake was estimated by using a food-frequency questionnaire. Self-reported HF was ascertained by using annual follow-up questionnaires with validation in a subsample. We used conditional logistic (or Cox) regression to estimate multivariable-adjusted ORs (or HRs) for HF and CAD. Results: Multivariable-adjusted ORs (95% CIs) for HF across consecutive quintiles of plasma trans 18:2 (linoleic acid) fatty acids were 1.0 (reference), 1.10 (0.79, 1.54), 0.88 (0.62, 1.25), 0.71 (0.49, 1.02), and 0.67 (0.45, 0.98) (P-trend = 0.01). Each SD of plasma trans 18:2 was associated with a 22% lower risk of HF (95% CI: 6%, 36%). Plasma trans 16:1 and 18:1 were not associated with risk of HF (P > 0.05). Dietary trans fats were not associated with incident HF or CAD. Conclusions: Our data are consistent with a lower risk of HF with higher concentrations of plasma trans 18:2 but not with trans 16:1 or trans 18:1 fatty acids in male physicians. Dietary TFAs were not related to incident HF or CAD.