A comparative study of dual organelle pH tracker, red-blue and red-green sensors and its applications in cancer cells studies

Abstract A systematic study on structurally similar, spectrally distinguishable dual-mode pH sensors ( 1 ) with red–blue and ( 2 ) with red–green emission is presented covering optical spectroscopic measurements in solution and in HeLa cells and their sensing performance is compared. Both sensors, which contain a piperazine unit that targets organelles such as lysosomes and mitochondria, exhibit different colors in acidic and neutral media, and rely on specific photophysical mechanisms for acidic pH sensing—namely photo-induced electron transfer (PET), intramolecular charge transfer (ICT), and fluorescence resonance energy transfer (FRET)—for acidic pH sensing. As determined from their emission and excitation spectra, dual-mode sensors 1 and 2 exhibit a broad range of p K a values, including four different p K a for 2 calculated by the ratiometric emission intensities in the neutral to acidic pH range. A comparison of the sensing behavior of these novel sensor molecules with Mito and Lysotracker dyes highlight their significant application potential, as the fluorescence of one fluorophore confirms the sensor’s presence within cells, while the other reflects the pH of the cellular environment.