This study was conducted to determine the association between place of residence (grouped into neighbourhoods), and 25-hydroxy D (25[OH]D) levels of individuals of Karachi. Addresses of 4788 individuals tested for 25[OH]D at the clinical laboratory of the Aga Khan University (AKU), Karachi, from January 2007 to June 2008 were reviewed. The neighbourhoods were categorized into ten, based on locality attributes. A high overall prevalence (74%) of vitamin D deficiency (VDD) was observed. There was a significant difference (p-value < 0.01) between mean log 25[OH]D levels amongst neighbourhoods grouped according to distinct housing structure attributes and localities. A high frequency of VDD in all the studied localities of an urban city warrant dietary vitamin D supplementation and food fortification.