Biocoat poly d lysine 96 well plates

For cytosolic pH measurement, cells were plated in quadruplicates in Corning BioCoat Poly-D-Lysine 96-well plates (Corning, 354640) at plating densities of 3 ×10⁴ cells perwell for HEK-293 cells and 1 ×10⁴ cells per well for UOK262 FH −/− and UOK262 FH rescue cells. Cells were allowed to adhere for 24 h and incubated with 10 µM of pHrodo™ green AM (Life Technologies, P35373) intracellular pH indicator at 37 °C for 30 mins as per manufacturer’s protocol. Standard curves of cytosolic pH were generated using pHrodo™ green AM intracellular pH calibration buffer kit (Life Technologies, P35379) that contains buffers at pH 4.5, 5.5, 6.5 and 7.5, as well as valinomycin and nigericin to help equilibriate the pH inside and outside the cells. The fluorescence was then measured using a Biotek Synergy MX plate reader, with pHrodo™ green AM detected using λ_ex of 509 nm and λ_em of 533 nm as per the manufacturer’s instructions.