BACKGROUND AND PURPOSE: Although studies have linked types of fatty acids with coronary heart disease, data on individual fatty acids and risk of ischemic stroke are limited. We aimed to examine the associations between serum fatty acid concentrations and incidence of ischemic stroke and its subtypes. METHODS: We conducted a prospective case-control study nested in the Women's Health Initiative Observational Study cohort of postmenopausal US women aged 50 to 79 years. Between 1993 and 2003, incident cases of ischemic stroke were matched 1:1 to controls on age, race, and length of follow-up (964 matched pairs). Conditional logistic regression was used to estimate odds ratios and 99.9% confidence intervals (CI) for ischemic stroke and its subtypes. RESULTS: The multivariable-adjusted odds ratios and 99.9% CI of ischemic stroke associated with a 1-SD increment in serum fatty acid concentration were 1.38 (99.9% CI, 1.05-1.83) for linoelaidic acid (18:2tt, SD=0.04%), 1.27 (99.9% CI, 1.06-1.51) for palmitic acid (16:0, SD=2.74%), 1.20 (99.9% CI, 1.01-1.43) for oleic acid (18:1n9, SD=2.32%), 0.72 (99.9% CI, 0.59-0.87) for docosapentaenoic acid (22:5n3, SD=0.18%), 0.72 (99.9% CI, 0.59-0.87) for docosahexaenoic acid (22:6n3, SD=0.91%), and 0.81 (99.9% CI, 0.67-0.98) for arachidonic acid (20:4n6, SD=2.02%). These associations were generally consistent for atherothrombotic and lacunar stroke but not cardioembolic stroke. CONCLUSIONS: These findings suggest that individual serum trans, saturated, and monounsaturated fatty acids are positively associated with particular ischemic stroke subtypes, whereas individual n3 and n6 polyunsaturated fatty acids are inversely associated.