BACKGROUND AND PURPOSE: Liver disease, particularly non-alcoholic fatty liver disease (NAFLD), is a risk factor for cardiovascular disease, but little is known about its relationship with ischemic stroke. METHODS: In the Reasons for Geographic and Racial Differences in Stroke (REGARDS) cohort of 30,239 American black and white adults, we assessed baseline NAFLD as fatty liver index (FLI) >60, and assessed liver biomarkers aspartate aminotransferase (AST), alanine aminotransferase (ALT), γ-glutamyl transpeptidase (GGT), and the AST/ALT ratio and risk of incident ischemic stroke over 5.8 years using a case-cohort study design. RESULTS: Considering 572 strokes and a 1,017-person cohort sample, NAFLD was inversely associated with stroke risk in men (HR: 0.50; 95% CI: 0.26, 0.96), as was being in the highest ALT quintile versus the lowest (HR: 0.39; 95% CI: 0.19, 0.78) and the highest versus lowest GGT quintile (HR: 0.45, 95% CI: 0.24, 0.85), but not in women. Conversely, FLI score above the 90th percentile was associated with increased stroke risk among women (HR: 2.26; 95% CI: 1.14-4.47), but not men. AST was not associated with stroke risk in either sex. AST/ALT ratio >2 was strongly associated with increased stroke risk in whites, but not blacks (HRs: 3.64; 95% CI: 1.42-9.35 and 0.97; 95% CI: 0.45-1.99, respectively; p for interaction = 0.03). CONCLUSIONS: The relationships between NAFLD, liver biomarkers, and ischemic stroke are complex, and sex and race differences we observed require further study and confirmation.