4.2. Vesicle Preparation

Large unilamellar vesicles (LUVs) were used as the membrane model in the infrared spectroscopy experiments. LUVs were prepared by the extrusion method with the mini-extruder from Avanti Polar Lipids. Appropriate amounts of POPC and POPG were weighed in a dark glass vial and dissolved in CHCl₃/CH₃OH 1/1 v/v. The organic solvent was removed under a gentle stream of nitrogen and the obtained lipid film was placed under vacuum pumping overnight. PBS was used to rehydrate the film, and after five vortex cycles of 1 min separated by 1 min rest, a suspension of large multilamellar vesicles was obtained. Then, five freeze–thaw cycles were performed by placing the vial in liquid N₂ and in a 323 K water bath. Finally, extrusion was carried out by passing the vesicle suspension (1 mL) 11 times through a polycarbonate filter (Whatman). In particular, the first extrusion was performed with a 400 nm pore size filter, then with a second 100 nm filter. To mimic the net charge of MOM,⁴⁰ a composition of 20%_mol negatively charged POPG and 80%_mol zwitterionic POPC was chosen.