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Abstract

Liquid-liquid phase separation in giant unilamellar vesicles (GUVs) leads to the formation of intramembrane domains. To mimic charged biological membranes, we studied phase separation and domain formation in \GUVs\} of ternary lipid mixtures composed of egg sphingomyelin, cholesterol, and the negatively charged lipid dioleoylphosphatidylglycerol. The \{GUVs\} were exposed to solutions of sucrose and high-saline buffer. The phase diagram was determined using epifluorescence microscopy for vesicle populations with symmetric and asymmetric solution compositions across the membranes. Trans-membrane solution asymmetry was found to affect the membrane phase state. Furthermore, compared to the case of salt-free conditions, the phase diagram in the presence of high-saline buffer (both symmetrically or asymmetrically present across the membrane) was found to exhibit a significantly extended region of liquid-ordered and liquid-disordered coexistence. These observations were confirmed on single \{GUVs\ using microfluidics and confocal microscopy. Moreover, we found that the miscibility temperatures markedly increased for vesicles in the presence of symmetric and asymmetric salt solutions. Our results demonstrate a substantial effect of salt and solution asymmetry on the phase behavior of charged membranes, which has direct implications for protein adsorption onto these membranes and for the repartitioning of proteins within the membrane domains.