Background and Purpose - Variations in the - 397T > C ( rs2234693) and - 351A > G ( rs9340799) single nucleotide polymorphisms of the estrogen alpha receptor ( ESR1) gene were found to be strongly associated with risk of ischemic heart disease, although not all studies could replicate this finding. One study also reported an association with stroke. We assessed whether variations in the ESR1 gene are associated with the risk of stroke in the general population. Methods - This prospective population- based study was based on 6229 Rotterdam Study participants who at baseline ( 1990 - 1993) were aged 55 years or older, free from stroke, and had assessment of the ESR1 rs2234693 and rs9340799 single nucleotide polymorphisms. Follow- up for incident stroke was complete until January 1, 2005. Data were analyzed with Cox proportional hazards models for men and women separately with adjustment for age. Results - During an average follow- up time of 10.1 years, 659 strokes occurred, of which 386 were ischemic. Three common haplotypes were identified: - 397T/ - 351A ( carried by 78% of all participants), - 397C/ - 351G ( carried by 57%), and - 397C/ - 351A ( carried by 22%). Although we had at least 89% power to detect a relative risk of 1.5 ( alpha = 0.05) in all subgroups, we did not find any association between ESR1 haplotype carriership and risk of stroke and ischemic stroke. Conclusions - We have not been able to replicate the previously reported association between variations in the ESR1 gene and risk of stroke.