Global energy consumption has increased tremendously throughout these recent years. Fossil fuels remain as our main source of energy despite being non-renewable and finite resource. This situation will eventually lead to shortage of energy if not addressed properly. In light of this issue, renewable energy has been one of the most promising alternatives to fossil fuels as energy source. Energy are not necessarily required at large scales. For instance, conventional batteries are widely used to power up small electrical appliances. However, these batteries are not without their drawbacks hence various researches have been conducted to develop fuel cells that can be used to power up small electrical appliances. This study aims to develop reference data by using glucose as substrate in the microbial fuel cell (MFC) as well as to screen fruit waste of orange, banana and mango to be used as substrate in the MFC. The reference data was successfully developed at different amount of glucose and different temperature. High glucose concentration gives higher peak voltage output compared to low glucose concentration. However, the voltage output at low glucose level shows more stable reading. When operated at room temperature of 28 ºC, the MFC produced highest peak voltage output at 360 mV. Meanwhile, the MFC of temperatures 40 °C, 50 °C and 60 °C produced highest voltage output of 259 mV, 255 mV and 320 mV respectively. Based on the collected data and other references, the MFCs were able to be effectively operated between 28 ºC and 60 °C. The results from screening of fruit wastes as substrate for MFC showed that fruit waste from orange produced highest voltage output 357 mV, followed by banana and mango fruit waste. © 2021 American Institute of Physics Inc.. All rights reserved.