SPICE‐Met: profiling and imaging energy metabolism at the single‐cell level using a fluorescent reporter mouse

The regulation of cellular energy metabolism is central to most physiological and pathophysiological processes. However, most current methods have limited ability to functionally probe metabolic pathways in individual cells. Here, we describe SPICE‐Met (Single‐cell Profiling and Imaging of Cell Energy Metabolism), a method for profiling energy metabolism in single cells using flow cytometry or imaging. We generated a transgenic mouse expressing PercevalHR, a fluorescent reporter for cellular ATP:ADP ratio. Modulation of PercevalHR fluorescence with metabolic inhibitors was used to infer the dependence of energy metabolism on oxidative phosphorylation and glycolysis in defined cell populations identified by flow cytometry. We applied SPICE‐Met to analyze T‐cell memory development during vaccination. Finally, we used SPICE‐Met in combination with real‐time imaging to dissect the heterogeneity and plasticity of energy metabolism in single macrophages ex vivo and identify three distinct metabolic patterns. Functional probing of energy metabolism with single‐cell resolution should greatly facilitate the study of immunometabolism at a steady state, during disease pathogenesis or in response to therapy. Synopsis The fluorescent sensor PercevalHR allows for measurements of cellular ATP:ADP ratio. Here, transgenic mouse expression of PercevalHR in conjunction with metabolic inhibitions enable the profiling of energy metabolism in cell populations or in single cells using flow cytometry or imaging. Modulation of PercevalHR fluorescence using metabolic inhibitors allows to infer cell dependencies on oxidative phosphorylation and glucose. Using flow cytometry, SPICE‐Met can simultaneously profile energy metabolism in complex cell populations simultaneously. SPICE‐Met and single cell imaging leads to the identification of three distinct energy metabolic profiles in peritoneal macrophage, thus revealing their metabolic heterogeneity and plasticity. Graphical Abstract A new transgenic mouse model allows analysis of energy metabolism at single‐cell resolution in living cells and is applied here to T cells and perinatal macrophages.