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Background Mean platelet volume (MPV) represents the platelet size, and is a surrogate marker of activated platelets. We investigated the relation between atrial fibrillation (AF) and MPV in patients with acute ischemic stroke. Methods Total 357 consecutive patients (mean age, 65.8±13.0 years; 37.8% women) with acute ischemic stroke within 72 hours of onset, were retrospectively enrolled. Patients were divided into two groups: stroke with AF and stroke without AF. Various parameters were analyzed and compared between the two groups. Results Sixty-three patients (17.65%) were enrolled in the stroke with AF group. The frequency of female gender, pre-existing coronary artery disease, heart failure and hyperlipidemia, age, and the NIHSS score at admission were all significantly higher in the stroke with AF group (P<0.05). The estimated glomerular filtration rate and triglyceride levels were significantly lower in the stroke with AF group (P <0.001 and P =0.001, respectively). MPV of stroke with AF group was significantly higher than that observed in stroke without AF group (8.4±1.0 and 8.0±0.9, respectively, P<0.01). The optimal cut-off value of MPV for distinguishing stroke with AF from stroke without AF was 7.95 (sensitivity 0.63, specificity 0.56, area under the curve 0.63). Multivariate logistic regression analysis demonstrated that MPV (odds ratio [OR], 1.52; P<0.05), previous anticoagulant medication (OR, 4.91; P<0.05), and NIHSS score on admission (OR, 1.14; P<0.001), were independently associated to stroke with AF group. Conclusions MPV increases with AF in acute ischemic stroke.