Abstract: Disclosed herein is a protein-based, genetically-encoded bioluminescent sensor that can report changes in intracellular ATP in live cells. This sensor is built from the NanoLuc luciferase (Promega) and mScarlet red fluorescent protein (DOI: 10.1038/nmeth.4074) in a novel platform that provides a much more quantitative optical signal for measurements. Analytical Chemistry demonstrates its characterization and that it can be used to image ATP differences in live cells under the skin of mice. This new sensor can be very useful for research purposes in cancer biology to measure cell health and chemotherapeutic drug efficacy for example in animal models of tumors or in cell-based screening.