In this work, a comprehensive experimental analysis on the performance of a 50 cm2 polymer electrolyte membrane (PEM) fuel cell is presented, including experimental results for a dedicated load cycling test. The harmonized testing protocols defined by the Joint Research Centre (JRC) of the European Commission for automotive applications were followed. With respect to a reference conditions representative of automotive applications, the impact of variations in the cell temperature, reactants pressure, and cathode stoichiometry was analyzed. The results showed that a higher temperature resulted in an increase in cell performance. A higher operating pressure also resulted in higher cell voltages. Higher cathode stoichiometry values negatively affected the cell performance, as relatively dry air was supplied, thus promoting the dry-out of the cell. However, a too low stoichiometry caused a sudden drop in the cell voltage at higher current densities, and also caused significant cell voltage oscillations. No significant cell degradation was observed after the load cycling tests.