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Objectives: There is no study about correlation between vitamin D serum level and multiple sclerosis (MS) in Iran. So in this survey, we investigated the serum level of 25-hydroxy vitamin D in MS patients and compared it with controls in central region of Iran, an area with medium to high risk for MS disease, in spite of high sun exposure., Methods: A case-control study was conducted from July 1, 2008 to July 31, 2009. We enrolled 50 definitive MS patients, according to McDonald's criteria as case group and 50 matched controls. Our age limits were 15 to 55 years and those patients with estimated expanded disability status scale less than 5 were introduced to the study. We measured serum level of 25-hydroxy vitamin D and compared them between the two groups., Results: We gathered 42 females and 8 males as case group and the same numbers as control group without significant age difference. The mean serum level of 25-hydroxy vitamin D in case and control groups were 48 and 62 nmol/L, respectively, and the difference was significant (P=0.036). Also, our study showed significant discrepancy between the two groups according to the rate of deficiency, insufficiency and normal range of vitamin D categories (P=0.021)., Conclusions: We found the same results as those studies carried out in Europe and North America; i.e., lower serum vitamin D level in MS patients than that in normal population, in spite of sufficient sun exposure in Isfahan region.