In this work, we present a feasibility study of the simultaneous use of electrochemical impedance spectroscopy (EIS) and synchrotron X-ray radiography in a polymer electrolyte membrane fuel cell (PEMFC) for studying the relationship between the presence of liquid water and cell performance. X-ray radiography was used to quantify liquid water in the anode and cathode fibrous carbon paper substrate and microporous layer (MPL). EIS was used as a complementary diagnostic tool to quantify the equivalent resistances of the fuel cell. Two fuel cell cases with varying MPL thicknesses were compared under constant current density operation. The fuel cell with a 150 μm-thick MPL resulted in an equivalent mass transport resistance that was 13% lower than the fuel cell with a 100 μm-thick MPL. This notable difference in mass transport resistance was attributed to the smaller quantity of liquid water in the cathode substrate, particularly in the transition region between the MPL and carbon paper. •PEM Fuel cell performance with 100 and 150 μm-thick MPLs were compared.•EIS and synchrotron X-ray radiography were used in tandem to diagnose PEMFCs.•Effect of MPL thickness on liquid water was visualized with synchrotron X-rays.•Liquid water in the cathode substrate was correlated to mass transport resistance.