A Polarity-Sensitive FLIM Probe With Large Lifetime Dynamic Range for Visualizing Intracellular Lipid Droplets.

Fluorescence lifetime imaging microscopy (FLIM) is a powerful tool for probing intracellular microenvironments, but successful polarity imaging relies on probes that combine high organelle specificity, pronounced lifetime responses, and sufficient sensitivity to detect subtle polarity variations in lipid droplets (LDs). In this study, we report BTDO-MEA, a fluorescent probe with a donor-acceptor structure for high-resolution imaging of intracellular LD polarity. BTDO-MEA shows excellent lipophilicity, a large Stokes shift, and a large linear fluorescence lifetime response to environmental polarity. We further demonstrated BTDO-MEA as a FLIM probe to visualize polarity changes of LDs in living cells. The probe enables discrimination the polarity differences between the core and the periphery of intracellular LDs. Upon oleic acid treatment, the overall lifetime of the LDs increased from 1.5 to 3 ns, indicating a significant polarity decrease in the periphery the LDs during the acute lipid accumulation process. These results demonstrate BTDO-MEA could be used as a useful imaging tool for monitoring lipid accumulation and studying lipid-associated metabolic disorders.