Using Synergy™ Mx Multimode Microplate Reader (Bio Tek Instruments), orange fluorescence was measured with an excitation wavelength of 480 nm and an emission wavelength of 590 nm, while green fluorescence was determined with an excitation wavelength of 480 nm and an emission wavelength of 530 (Figure 3). Automated Mito-ID™ Membrane Potential Cytotoxicity Procedure. Initial experiments used the compound carbonyl cyanide 3-chlorophenylhydrazone (CCCP).CCCP is a proton ionophore that destroys the membrane potential across the mitochondrial membrane.With mitochondrial damage or loss of membrane potential, the dye cannot accumulate in the mitochondria, which is indicated by a lack of orange fluorescence.Using a single excitation wavelength both monomers and aggregates can be measured with dual wavelength fluorescence emission determination (Figure 2). Schematic illustration of Mito-ID™ Membrane Potential dye aggregation in the mitochondria as a result of membrane polarization.
The EL406 Combination Washer Dispenser is well suited to the Mito-ID™ cytotoxicity assay workflow.
It offers fast, accurate media removal and plate washing capabilities through its Dual-Action™ Manifold.