Fully treated cells show hydrogenosomes with an electron-dense deposit which aggregates to a variable extent.The staining is seen in the interior of hydrogenosomes in some instances. It is also observed by microscopy that the K ⁺ /H ⁺ ionophor nigericin does not inhibit hydrogenosomal loading with BCECF.The pH-sensitive fluorescent dyes to measure cytosolic pH. 1.Prepare a 2 to 20 mM stock solution of BCECF in DMSO. 2.Prepare a 5-50 μM BCECF dye-loading solution in buffer solutions (HHBS or PBS).3. Add 1000 μL/well (6-well plate),100 μL/well (96-well plate) or 25 μL/well (384-well plate) BCECF dye-loading solution into the cell plate.4. Incubate the dye-loading plate in a cell incubator for 30-60 minutes.5. Wash and replace the dye-loading solution with buffers.6. Run the pH assay by monitoring the fluorescence at E _x /E _m = 490/535 nm or 430/535 nm for ratio measurements.