A bright red fluorescent genetically encoded sensor for lactate imaging

Lactate plays a crucial role in energy metabolism and greatly impacts protein activities, exerting diverse physiological and pathological effects. Therefore, convenient lactate assays for tracking spatiotemporal dynamics in living cells are desirable. In this paper, we engineered and optimized a red fluorescent protein sensor for l-lactate named FiLa-Red. This indicator exhibited a maximal fluorescence change of 730 % and an apparent dissociation constant (K_d) of approximately 460 μM. By utilizing FiLa-Red and other sensors, we monitored energy metabolism in a multiplex manner by simultaneously tracking lactate and NAD⁺/NADH abundance in the cytoplasm, nucleus, and mitochondria. The FiLa-Red sensor is expected to be a useful tool for performing metabolic analysis in vitro, in living cells and in vivo.