Lysosensor yellow blue dnd 160 dye

Lysosomal pH was measured using LysoSensor Yellow/Blue DND-160 dye (L7545; Thermo Fisher Scientific) according to^{76 (link)}.

Endosomal pH was determined using the LysoSensor Yellow/Blue DND-160 dye (L7545; Thermo Fisher Scientific). RPTEC/TERT1 cells were cultured in IBIDI chambers for 10 d. Cells were washed three times with isotonic solution containing 2.5 mM KCl, 140 mM NaCl, 1.2 mM CaCl₂, 0.5 mM MgCl₂, 5 mM glucose, and 10 mM Hepes (305 mosmol/litre, pH 7.4 adjusted with Tris), and then treated with LysoSensor for 10 min at 37°C. After the incubation period, cells were washed again with isotonic solution and imaged using Leica Thunder Imager 3D Cell Culture Microscope for 3 min without stimulation. In continuation, cells were treated with vehicle (DMSO) or 100 nM bafilomycin A1 to get a normalization point. Images were acquired with a 5X objective on Leica Thunder Imager 3D Cell Culture Microscope and processed using ImageJ software.