摘要：SummaryIn situmeasurement of cellular metabolites is still a challenge in biology. Conventional methods, such as mass spectrometry or fluorescence microscopy, would either destroy the sample or introduce strong perturbations to target molecules. Here, we present multiplex stimulated Raman scattering (SRS) imaging cytometry as a label-free single-cell analysis platform with chemical specificity and high-throughput capabilities. Using SRS imaging cytometry, we studied the metabolic responses of human pancreatic cancer cells under stress by starvation and chemotherapeutic drug treatments. We unveiled protrusions containing lipid droplets as a metabolic marker for stress-resistant cancer cells. Furthermore, by spectroscopic SRS mapping, we unveiled that triglyceride in lipid droplets are used for local energy production through lipolysis, autophagy, and β-oxidation. Our findings demonstrate the potential of targeting lipid metabolism for selective treatment of stress-resistant cancers. Collectively, these results highlight SRS imaging cytometry as a powerful label-free tool for biological discoveries with a high-throughput, high-content capacity.Graphical AbstractDisplay OmittedHighlights•Multiplex SRS imaging cytometry is developed for single-cell metabolic analysis•Lipid-rich protrusions are unveiled as a marker for stress-resistant cancer cells•Lipids in protrusions are used for local energy production via β-oxidation•Blockage of β-oxidation impairs survival of stress-resistant cancer cellsOptical Imaging; Metabolomics; Cancer
