Nanotechnology has offered enormous improvement in field of therapeutics by means of designing of nano-carriers that can cross biological barriers and are able to target cellular reservoirs. The aim of the present investigation was to formulate and evaluate Niosomes, a synthetic microscopic non-ionic surfactant vesicles formed on the admixture of a non-ionic surfactant, cholesterol and phosphate with subsequent hydration in aqueous media. The delivery of drugs by “vesicular drug delivery system” such as niosomes provides several important advantages over conventional drug therapy. Ofloxacin was selected as a suitable drug for the present study because it is a potent secondgeneration fluoroquinolone active against a broad range of gram positive and gram-negative aerobic and anaerobic bacteria. In the present investigation three formulations of Niosomal drug delivery system of ofloxacin with non ionic surfactant, span 60, in various proportions were prepared and evaluated for Morphological characterization, Encapsulation efficiency, In-vitro drug release study. The Ofloxacin niosomes were prepared by lipid film hydration method. The prepared ofloxacin niosomes showed a vesicle size of 100-300 nm, the entrapment efficiency was 78.4%. The partition coefficient value of ofloxacin was found to be 0.5.