Abstract : The constantly increasing interest in transdermal delivery drives directly from the advantages of the transdermal route compared with oral route. The aim of the study was to formulate, optimize and in vitro characterization of an effective ethosomal formulation of Satranidazole and to investigate the influence of different preparation conditions on the performance of the formulation to enhance the transdermal delivery. From the 9 formulations with varying concentration of ethanol (from 20%w/v to 40% w/v) and lecithin (from 1%w/w to 3% w/w), the highest entrapment efficiency of 97.1 % and good release obtained for F7 formulation. The 4 vesicles produced by different techniques were characterized for drug encapsulation efficiency, zeta potential, vesicular size and other morphological studies like SEM analysis. The vesicular size of the formulations varied from 200 nm – 400 nm, the optimized formulation showed vesicle size of 246.44 ±0.031 nm. The in vitro release kinetics of drug was found to be zero order .The in-vitro antibacterial studies against Staphylococcus aureus showed significant activity for ethosomal formulation than the control, the MIC was found to be 5?g/ml. The SEM analysis (Figure 7) of F7 formulation indicates that that the drug molecules were entrapped in the polymers .The stability studies of the optimized formulation at room temperature (30 ± 2°C) and at refrigerator temperature (4 ± 2°C) shows that the formulation is more stable at low temperature compared to the room temperature. The overall results suggest that a suitably developed ethosomal formulation of Satranidazole can be of actual value for improving its clinical effectiveness in transdermal delivery. This study pointed out the influence of the structure and the size of the vesicles and consequently the technique used for the preparation, on the drug entrapment efficiency and on permeation rate.