A method for an efficient formation of giant vesicles of neutral phospholipids in microarrays using a microplotter equipment

A methodology which provides a high efficiency of giant vesicle formation was established using the gentle hydration method and a microplotter equipment. The method consists of preparing a mixture of zwitterionic egg yolk phosphatidylcholine/additive in solution and printing a number of droplets onto a glass substrate, which immediately dry after deposition. Then, gentle hydration of these micro-sized thin-films provides a high amount of giant liposomes, per microsized film. Several cases were studied by varying different compounds as additives (i.e., non-electrolytes and electrolytes) at different molar ratios, lipid to additive, in order to find the optimal conditions. Optical and confocal microscopies were employed to characterize vesicle formation. Studies indicate that the kosmotropic salt KH2PO4 at 1:10 molar concentration, EggPC to salt, is the most effective in vesicle production. Abundant liposome formation can be observed in a short time, about 5 min upon hydration. The osmotic pressure is the driven force to produce giant liposomes in our experiments, which is generated by dissolving the additive among two lipid lamellar phases in water. In salt experiments, the osmotic pressure strength is manly determined from the ion-specificity effect (i.e., the Hofmeister effect) rather than the concentration of the salt. The use of a salt as additive provides giant unilamellar vesicles (GUVs). The microplotter protocol provides benefits such as a facile, efficient and rapid way to prepare GUVs in mild conditions (i.e., free of solvents).