Lysosomal pH Measurement

LysoSensor Yellow/Blue DND-160 (L7545; Invitrogen/Thermo Fisher Scientific) was used for pH measurement of lysosomes. Cells were labeled with 3 μmol/L LysoSensor Yellow/Blue DND-160 in medium for 5 minutes at 37°C, washed twice with PBS, and immediately analyzed by using a microplate reader (excitation, 360 nm; emission, 451 and 518 nm). Quantification of lysosomal pH involved a ratio (emission 451/emission 518) according to pH calibration. The protocol of LysoSensor pH calibration measurement was adapted from the manufacturer’s instructions.