Dimensions, stability, and deformability of DOPC-cholesterol giant unilamellar vesicles formed by droplet transfer

<ns3:p>Background Understanding cell membrane-like lipid bilayers is crucial for studying fundamental biological mechanisms. Giant Unilamellar Vesicles (GUVs) are key tools for this investigation and have applications in both synthetic biology and, more recently, in microrobotics. The effects of cholesterol, a key component of cellular membranes, on synthetic phospholipid membrane models like GUVs are however not fully understood, as they may vary with lipid composition and production method. Methods We examined the size distribution, temporal stability and deformability of GUVs prepared with the droplet transfer method using different Dioleoylphosphatidylcholine (DOPC) to cholesterol ratios in the oil phase (namely 100:0, 85:15, 71:29, 60:40). Phase-contrast microscopy assessed size and stability, while deformability was tested by loading the GUVs with an aqueous ferrofluid and applying a uniform magnetic field to induce their elongation. Image analysis was conducted using Fiji and a custom Julia script. Results The median diameters increased with the content of cholesterol, together with the dimensional distribution. In terms of stability, cholesterol generally reduced GUV median diameter over time, while it varyingly influenced the number of vesicles. As for deformability, beyond the expected elongation dependent on the intensity of the applied magnetic field, there were no statistically significant differences in GUV deformability in the presence or absence of cholesterol. Conclusions Our findings suggest that cholesterol can lead to increased average diameter of GUVs made with DOPC through droplet transfer, while varyingly affecting their time-stability and not affecting their deformability. This study shows how small adjustments on a straightforward protocol like the droplet transfer method, provide a simple and effective way of tailoring GUV properties. Edits in the oil phase enable precise tuning of GUV membranes providing a tool for both fundamental studies and applications such as artificial cells or microrobots.</ns3:p>