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Introduction: In 2012 a centralization and specialization of stroke services was implemented in Central Region Denmark (CRD) (n= 1.3 million inhabitants). It implied that acute stroke care was to be provided at only 2 units with re-vascularization therapy.Objective: The impacts on length of acute hospital stay (AHS), rate of thrombolysis (IV tPA), evidence-based clinical care and mortality.Methods: Population-based before-and-after registry study. The study cohort included all stroke cases in Denmark, with patients outside CRD being used as comparison to account for general changes in stroke care. The period before (May 2011- April 2012) was compared to after (May 2013 - April 2014) using regression methods, including difference-in-differences (DID) analysis. Potential confounders included age, gender, civil status, previous strokes, diabetes, atrial fibrillation, smoking, alcohol, stroke severity, hypertension and type of stroke.Results: Baseline data in Figure 1. Median length of AHS (days) in CRD decreased from 5 (IQR 7) to 2 (3) vs. from 5 (9) to 5 (8) in the rest of Denmark. IV tPA rates increased from 16% (95CI 14-17) to 19% (17-21) of all acute ischemic strokes in CRD and from 9% (8-10) to 14% (13-15) in the rest of Denmark (DID RR 0.77 (0.66-0.91)). All-or-none rates of 11 process performance measures of in-hospital care increased from 51% (49-53) to 63% (61-65) in CRD vs. 49% (48-50) to 60% (59-61) in the rest of Denmark (DID RR 0.99 (0.93-1.05)). Adjusted 30-days mortality rate decreased non-significantly and comparable to the rest of the country; OR 0.97 (0.71-1.32) vs. OR 0.91 (0.77-1.07) (DID OR 1.03 (0.75-1.41)).Conclusions: Centralization of acute stroke care was associated with a significant reduction in length of AHS when compared to the development in the rest of Denmark. The use of IV tPA and the quality of acute stroke care also improved, but the trend was not different from the rest of Denmark. No changes in the adjusted 30-days mortality were observed.