Direct ethanol fuel cells (DEFCs) have been paid more and more attention in recent years due to the following advantages of ethanol: low toxicity, renewability and easy production by the fermentation of agricultural products [1]. Based on the present status of the electrolyte development, Naifon® membrane series is the most commonly used. It is well known that its conductivity is dependent on the water content in the membrane in a more a less linear fashion [2]. This necessitates sufficient water in the polymer electrolyte. On the other hand, water content can not be so high that the electrode will be flooded. Proper management of water through the membrane is indispensable for the desirable cell performance. Furthermore, ethanol can also permeate across Nafion® membranes, leading to a mixed potential and catalyst poison at the cathode, consequently causing the decreased cell performance and fuel utilization [3]. These both issues make it necessary to investigate the ethanol/water permeation behavior through membrane electrode assembly in PEMFCs. In the present work, water and ethanol crossover rates through membrane electrode assembly (MEA) were determined in a polymer electrolyte fuel cell (PEMFC). The investigated MEA consisted of two Pt/C electrodes (40 wt. %) and a Nafion®-115 membrane as the electrolyte. The anode compartment was pumped by ethanol aqueous solutions with different concentrations, and the cathode was supplied with high-purity helium at different flow rates to sweep off the permeated ethanol and water. The effluent from the cathode was on-line determined by gas Chromatograph (GC-14B, Shimadzu). The effect of the operation parameters such as the cell temperature, the carrier gas (He) flowrate at the cathode and the concentration of ethanol aqueous solutions on the ethanol/water permeation behavior was investigated. Based on the experimental results, it was found that the crossover rates of both water and ethanol increase with the increment of the operation temperature and the carrier gas (He) flow rate at the cathode. It was also found that the ethanol concentration had a significant effect on the crossover rates of ethanol and water. In the case of ethanol, the permeation rates presented a volcano behavior as the ethanol concentration increased, reaching a peak value when the ethanol concentration was 6.0 mol/L, which could be attributable to the different swelling behavior of Nafion® membrane in different ethanol aqueous solutions