A Novel Heterocyclic Xanthene-Analogous pH Probe for ­Quantitative Monitoring of Cell Surface pH by Fluorescence ­Lifetime Imaging

Abstract

Rapidly capturing slight changes in cell surface pH is extremely important to evaluate the rapid diffusion of acidic metabolites into the extracellular environment caused by disease and physiological pH fluctuations of cells. In this work, we designed a membrane-targeted pH probe, Mem-C_OC₁₈ , based on a novel heterocyclic xanthene-analogous backbone. Mem-C_OC₁₈ shows specific and stable staining ability towards membrane. Importantly, the fluorescence lifetime of Mem-C_OC₁₈ is highly sensitive against acidity within membrane, which is in favor of quantifying pH through fluorescence lifetime imaging. Using Mem-C_OC₁₈ , we recorded pH changes of 0.61 units on the surface of human cervical cancer cells (Hela) during glycolysis. Further on, we observed a robust pH-regulating mechanism of the plasma membrane that the pH fluctuation range within membrane (5.32–6.85) is much smaller than the change in extracellular environment (4.00–8.00). Consequently, we demonstrate a pH probe for quantifying small pH fluctuations within cell membrane that merits further evaluation for biology applications.

Key words

FLIM - heterocyclic xanthene-analogous - plasma membrane targeted - cell surface pH - glycolysis

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