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Background: Vitamin D deficiency is present in India in epidemic proportions despite plenty of sunshine. Reduced plasma 25(OH) D concentrations as a diagnostic marker of vitamin D deficiency have been in past decade associated with several well-established risk factors for ischaemic stroke, such as arterial hypertension, thrombosis, atherosclerosis. The aims and objectives of this study was to compare the serum 25(OH) D levels between the first ever acute stroke patients and healthy controls. Methods: A cross-sectional, case control study was conducted in a tertiary care hospital in New Delhi situated in north India. Serum 25‑hydroxyvitamin D (25(OH) D) levels in 85 patients of ischemic stroke, presenting within 7 days of onset of stroke was measured and was compared with 70 age and gender matched controls. Results: The mean age was 61.02±11.58 years and 58.63±11.28 years in cases and controls respectively. Females constituted 37.6% of the total number of cases and 43.4% of the controls. The age and gender-distribution were comparable between the cases and controls. The median value (IQR) of serum 25(OH) vitamin D level was 7.94 ng/mL (4.59-14.00) in the cases and it was 8.82 ng/mL (5.59-14.70) in the controls. The difference between the serum 25(OH) vitamin D levels of the two groups was not found to be statistically significant. Conclusions: There is a high prevalence of biochemical hypo-vitaminosis D in apparently healthy Indians of all age and sex groups despite adequate sunshine. There is no association between low vitamin D levels and stroke.