Osmotic pressure enables high yield assembly of giant vesicles in solutions of physiological ionic strengths
Giant unilamellar vesicles (GUVs) are micrometer scale minimal cellular mimics that are useful for synthetic biology and drug delivery. Unlike assembly in low-salt solutions, assembly of GUVs in solutions with ionic concentrations of 100-150 mM Na/KCl (salty solutions) is challenging. Chemical compounds deposited on the substrate or in the lipid mixture could assist in the assembly of GUVs. Here, we investigate quantitatively the effects of temperature and chemical identity of six assisting polymeric and small molecule compounds on the molar yields of GUVs composed of three different lipid mixtures using high resolution confocal microscopy and large dataset image analysis. All the polymers moderately increased the yields of GUVs either at 22 or 37 degrees Celsius, whereas the small molecule compound was ineffective. Low gelling temperature agarose is the singular compound that consistently produces yields of GUVs of greater than 10 %. We propose a free energy model of budding to explain the effects of polymers in assisting the assembly of GUVs. The osmotic pressure exerted on the membranes by the dissolved polymer balances the increased adhesion between the membranes, thus reducing the free energy for bud formation. Data obtained by modulating the ionic strength and ion valency of the solution shows that the evolution of the yield of GUVs support our models prediction. In addition, polymer specific interactions with the substrate and the lipid mixture effects yields. The uncovered mechanistic insight provides a quantitative experimental and theoretical framework to guide future studies. Additionally, this work shows a facile means for obtaining GUVs in solutions of physiological ionic strengths.