Electroformation of Giant Unilamellar Vesicles: Investigating Vesicle Fusion versus Bulge Merging

Abstract

Partially ordered stacks of phospholipid bilayers on a flat substrate can be obtained by the evaporation of a spread droplet of phospholipid-in-chloroform solution. When exposed to an aqueous buffer, numerous micrometric buds populate the bilayers, grow in size over minutes, and eventually detach, forming the so-called liposomes or vesicles. While observation of vesicle growth from a hydrated lipid film under an optical microscope suggests numerous events of vesicle fusion, there is little experimental evidence for discriminating between merging of connected buds, i.e., a shape transformation that does not imply bilayer fusion and real membrane fusion. Here, we use electroformation to grow giant unilamellar vesicles (GUVs) from a stack of lipids in a buffer containing either (i) nanometric liposomes or (ii) previously prepared GUVs. By combining different fluorescent labels of the lipids in the substrate and in the solution, and by performing a fluorescence analysis of the resulting GUVs, we clearly demonstrate that merging of bulges is the essential pathway for vesicle growth in electroformation.

Translocation of silica nanospheres through giant unilamellar vesicles (GUVs) induced by a high frequency electromagnetic field

“…The main advantages of this single layer lipid vesicle membrane system include ease of preparation and handling and close resemblance to the basic compartment structure of biological cells. 34,35 The GUVs used in this study are composed of conventional phospholipids with one hydrophilic head group attached to two lipophilic chains 35 that are directed towards the aqueous media and hydrophobic fatty acid chains forming the interior layer of the bilayer. 35,36 The formation of liposomes occurred in less than 5 min aer the aqueous sugar solution was added to the partially ordered stack of lipids due to spontaneous swelling of the dry lipid lms, with the hydration process le undisturbed throughout the procedure.…”

Production of Isolated Giant Unilamellar Vesicles under High Salt Concentrations

“…Due to the dense packing and the influence of the electric field, the membrane buds fuse which can be observed during the formation process. Recently, it has been shown that enclosed and detached vesicles do not fuse during electroformation (Micheletto et al, 2016 ). By decreasing the frequency of the AC field, the swollen vesicles follow the slow change of the field and detach from the surface, forming free floating GUVs (Politano et al, 2010 ).…”

Giant Lipid Vesicle Formation Using Vapor-Deposited Charged Porous Polymers

“…We surmise that polymer substrates resulting in high vesicle yields (such as the 36 μm thick membrane deposited at −20 °C) also result in larger vesicles; this is likely due to coalescence when many vesicles are present in a given area. Similar to vesicle bulge merging in electroformation, vesicle coalescence via merging through the surface of polymer has been shown for both agarose and dextran PEG hydrogels. , …”