Proton conduction and oxygen transport in the cathodes of direct methanol fuel cells (DMFCs) are affected by the loading and distribution of Nafion ionomer. These effects of ionomer in the catalyst layer with carbon-supported catalysts on the cathode performance were investigated by electrochemical techniques and single cell testing in order to improve the power output of DMFC. It is found that varying the Nafion content has minimal effect on the electrochemical surface area (ESA) and catalyst utilization, but has significant effect on pore structure, proton conduction, and oxygen transport in the catalyst layer. A higher content of Nafion ionomer is needed in DMFC cathodes to achieve the same specific conductivity as in proton exchange membrane fuel cells (PEMFC) due to the higher catalyst loading in DMFCs. Optimal cell performance is achieved with a balance among catalytic activity, proton conductivity, and oxygen transport. These findings could provide guidelines for electrode design, fabrication, and prediction of cell performance. Copyright (C) 2012, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.