Giant unilamellar vesicles (GUVs), cell-sized membrane compartments, have proven to be useful research tools for many biomembrane studies including lipid membrane organization, lipid dynamics, membrane division and fusion, and membrane protein function to name a few. Previously, we developed a new solvent-free method to produce GUVs using nanocellulose paper termed PAPYRUS for Paper-Abetted liPid hYdration in aqUeous Solutions. To date, solvent-free techniques used to obtain GUVs in physiological concentrations of salts often result in low yields or rely on the dissolution of molecules into solution. In this work, we show that high yields of GUVs, O(10<sup>6</sup>) per µg lipid, can efficiently be obtained in physiological conditions through a simple modulation of the ionic strength of the solution. After first allowing a period of growth in low salt conditions, the modulation is carried out by introducing a concentrated amount of salt to the sample. Using a quantitative confocal microscopy based methodology, we show that increasing the salt concentration from 0.01 mM to 150 mM results in a 30-fold decrease in the GUV yields produced from nanocellulose paper. We subsequently show the full recovery of the GUV yields by increasing the modulation time from 0 to 10 minutes. As an example of the utility of the modulated PAPYRUS technique for creating synthetic cell-like structures, we assemble GUVs with an actin cytoskeleton. This work increases our understanding of the effects of ionic strength on the process of assembly of GUVs. It also opens new avenues for assembling large quantities of GUVs useful for biomembrane research and applications.

High yields of giant vesicles in physiological conditions by a modulation of ionic strength

ACS Spring Meeting 2021

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