In PEM fuel cells, one of the most important components is a gas diffusion layer (GDL). The GDL performs the essential functions such as passage for reactant gases from flow field channels to catalyst layers, removal of produce water from the catalyst layer area to flow field channels, electronic conductivity by providing passage for electron traort from bipolar plates to catalyst layers, heat conductivity by providing efficient heat conduction between bipolar plates and membrane electrode assembly (MEA), mechanical support to the membrane electrode assembly (MEA), and protection of the catalyst layer from corrosion or erosion caused by flows or other factors. The GDL plays a crucial role in water management that maintains the delicate balance between membrane hydration and water removal from cell. GDL manufacturing process is an effective parameter on PEM fuel cell performance, because it can change the structure and composition of the GDL. In this study, electrochemical techniques were applied to investigate the effect of four cathode diffusion layer inks mixing procedures on the microstructure of gas diffusion electrode and polymer electrolyte fuel cell performance. Homogeneity of gas diffusion layer inks have been supplied following four methods: three techniques of using ultrasound waves including pulse ultrasonic, continuing ultrasonic and bath ultrasonic, and a technique of using mechanical force which was magnetic stirring. GDL inks for cathode electrodes had the same compositions but they prepared by different mentioned techniques. The performance of cell with the electrode made by pulse ultrasonic technique was also measured under different operational conditions such as various cell temperatures, pressures and cathode relative humidity in order to choose proper operational condition for comparative tests. Then, four electrodes which had diffusion layers made by different techniques were evaluated at chosen operational condition. The performance of cells was evaluated by electrochemical tests such as polarization curves and electrochemical impedance spectroscopy. Lastly, morphological images of the GDLs in MEA have been analyzed by scanning electron microscopy (SEM). Results of the cell performance showed an improvement in maximum power density in order of 30% for the cell that its diffusion layer was made by bath ultrasonic technique in comparison to other cells. Electrochemical impedance spectroscopy confirmed the polarization results. This can be attributed to the effect of manufacturing process on the homogeneity and porosity of diffusion layer microstructure. SEM micrographs showed that the type of diffusion layer mixing method can influence on its surface morphology. Surface morphology of diffusion layer made by bath ultrasonic technique revealed that it has a more homogenous and crack free microstructure in comparison to surface morphology of other diffusion layers. The diffusion layer made by magnetic stirring had a non homogenous surface with irregular channels.