This video describes a confocal microscopy imaging-based assay to measure mitochondrial calcium uptake in cultured cells. The assay uses the calcium-sensitive fluorescent dye Rhod-2/AM to measure the mitochondrial calcium in permeabilized cultured cells.
Protocol
1. Reagents and Solutions NOTE: The second part is experimental procedure for confocal imaging of mitochondrial Ca2+ in cultured cells Make Tyrode's solution (130 mM NaCl, 4 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 10 mM glucose, and 10 mM HEPES, pH 7.2 with KOH). Store it at 4 °C. Warm it to room temperature before use. Prepare Wash Solution (100 mM potassium acetate, 15 mM KCl, 5 mM KH2PO4, 5 mM Mg-…