Giant membrane vesicles (GUVs) and Giant plasma membrane vesicles (GPMVs) are used as models to study membrane properties. We conducted a comparative study to examine how reducing the volume of vesicles with different lipid compositions, solution symmetries, solution asymmetries, and membrane charges affects their morphology. We used three‐dimensional visualization techniques to study the shape of the vesicles. Although the vesicles may not be perfectly spherical, they exhibit some fluctuations in their shape. To understand these variations, we used confocal image stacks for visualization. Our experimental observations show that the membrane's charge influences the deflation of the GUVs in the presence of trans‐bilayer sugar asymmetries. The lipid bilayers of our GUVs have a uniform distribution of lipids in both leaflets, indicating no asymmetry in lipid composition. However, we induce trans‐bilayer asymmetries by exposing each leaflet to different solution compositions, leading to asymmetric adsorption or desorption layers. We also estimated and compared the deformation of GPMV shapes extracted from cells with trans‐bilayer buffer asymmetries and composition asymmetries with biomimetic membranes.
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