As a new form of regulated cell death, ferroptosis is closely related to various diseases. To interpret this biological behavior and monitor related pathological processes, it is necessary to develop appropriate detection strategies and tools. Considering that ferroptosis is featured with remarkable lipid peroxidation of various cell membranes, it is logical to detect membranes' structural and environmental changes for the direct assessment of ferroptosis. For this sake, we designed novel polarity-sensitive fluorescent probes Mem-C₁C₁₈ and Mem-C₁₈C₁₈, which have superior plasma membrane anchorage, high brightness, and sensitive responses to environmental polarity by changing their fluorescence lifetimes. Mem-C₁C₁₈ with much less tendency to aggregate than Mem-C₁₈C₁₈ outperformed the latter in high resolution fluorescence labeling of artificial vesicle membranes and plasma membranes of live cells. Thus, Mem-C₁C₁₈ was selected to monitor plasma membranes damaged along ferroptosis process for the first time, in combination with the technique of fluorescence lifetime imaging (FLIM). After treating HeLa cells with Erastin, a typical ferroptosis inducer, the mean fluorescence lifetime of Mem-C₁C₁₈ displayed a considerable increase from 3.00 to 4.93 ns, with a 64% increase (corresponding to the polarity parameter Δf increased from 0.213 to 0.232). Therefore, our idea to utilize a probe to quantitate the changes in polarity of plasma membranes proves to be an effective method in the evaluation of the ferroptosis process.

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