Ultrasensitive determination of intracellular superoxide in individual HepG2 cells by microfluidic chip electrophoresis

A microchip electrophoresis method was established for the determination of intracellular superoxide (O2−) in individual HepG2 cells. Dihydroethidium (DHE) was used as the specific fluorescent probe to react with intracellular O2− to form the fluorescent 2-hydroxyethidium. Excellent resolution between 2-hydroxyethidium and ethidium cation (E+) can be achieved within 20 s. E+ was reported to be generated from photochemical oxidation of DHE and interfere the determination of O2− with fluorescence microscopic technique. An extremely low detection limit of 2.0 amol was achieved owing to the minute sample volume and insignificant dispersion effect during microfluidic chip-based electrophoretic separation. Furthermore, only 2-hydroxyethidium peak was detected with the suggested single-cell analysis method, which indicates the photooxidation of DHE to E+ could be blocked by isolating either oxygen or light from them.