This project was initiated to evaluate the effect of alternative process temperatures to 38 °C at the anaerobic digestion step in a Swedish wastewater treatment plant (WWTP) treating mixed sludge. The efficiency of the different temperatures was evaluated with respect to biogas production, volume of sludge produced and nutrient content in the reject water to find the optimum temperature for the WWTP as a whole. Three temperatures, 34 °C, 38 °C and 42 °C, were compared in laboratory scale. Increasing the process temperature to 42 °C resulted in process instability, reduced methane yield, accumulation of volatile fatty acids and higher treatment costs of the reject water. By decreasing the temperature to 34 °C, slightly higher sludge mass was observed and a lower gas production rate, while the specific methane produced remained unchanged compared to 38 °C but foaming was observed at several occasions. In summary 38 °C was proved to be the most favourable temperature for the anaerobic digestion process treating mixed sludge when the evaluation included effects such as foaming, sludge mass and quality of the reject water.