Liposomes were prepared from 1,2-dioloeyl-sn-glycero-3-phosphocholine (DOPC, Avanti Polar Lipids, Inc.) with and without bovine heart CL (Avanti Polar Lipids). DOPC (6 mM) was dissolved in 1 mL CH2Cl2 with and without CL (6 mM) at room temperature.
After evaporation of the DOPC solution with an evaporator, 1 mL of 25 mM HEPES buffer, pH 7.4, was added to the precipitate at room temperature. The solution was shaken with a vortex for 1-2 min at room temperature, and ultrasonicated for 1-2 min at 25 °C.
The ultrasonicated solution was frozen with liquid nitrogen and incubated at 50 °C until the sample was melted. The shaking, ultrasonic, freezing, and melting were repeated three times. Liposomes were added to the protein solutions at room temperature.
After evaporation of the DOPC solution with an evaporator, 1 mL of 25 mM HEPES buffer, pH 7.4, was added to the precipitate at room temperature. The solution was shaken with a vortex for 1-2 min at room temperature, and ultrasonicated for 1-2 min at 25 °C.
The ultrasonicated solution was frozen with liquid nitrogen and incubated at 50 °C until the sample was melted. The shaking, ultrasonic, freezing, and melting were repeated three times. Liposomes were added to the protein solutions at room temperature.