The aim of the present study was to investigate the potential of transfersomal formulations for transdermal delivery of Stavudine and to evaluate Span 80, Tween 80, Sodium cholate and Sodium deoxycholate as edge activators. Vesicles containing phosphatidylcholine (PC) mixed with edge activators and Stavudine were prepared by conventional rotary evaporation method and characterized by various parameters (vesicles shape and surface morphology, size and size distribution, entrapment efficiency, elasticity, turbidity and in vitro drug release). The compositions of refined formulations were predicted, liposomes prepared and tested against control. Skin permeation profile of transfersomal formulation bearing Stavudine was observed and the investigations revealed an enhanced transdermal flux (8.91±0.9 μg/cm2/hr) and decrease lag time (0.9 hr) for Stavudine. The obtained flux was nearly 7.5 and 12.04 times higher than conventional liposomal formulation bearing Stavudine and plain drug solution. These results suggested that transfersomes are potential vehicles for improved transdermal delivery of Stavudine and Span 80, Tween 80 and Sodium cholate were equivalent to Sodium deoxycholate as edge-activators.