The blue cathode buffer (50 mM Tricine, 15 mM BisTris, 0.02% Coomassie Blue G-250 (pH 7.0) at 4 °C) was changed to the colorless cathode buffer (50 mM Tricine, 15 mM BisTris (pH 7.0) at 4 °C) once the running front reached halfway through the separating gel.