Micron-sized domains in quasi single-component giant vesicles

Giant unilamellar vesicles (GUVs), are a convenient tool to study membrane-bound processes using optical microscopy. An increasing number of studies highlights the potential of these model membranes when addressing questions in membrane biophysics and cell-biology. Among them, phase transitions and domain formation, dynamics and stability in raft-like mixtures are probably some of the most intensively investigated. In doing so, many research teams rely on standard protocols for GUV preparation and handling involving the use of sugars. Here, we demonstrate that following such a standard approach can lead to abnormal formation of micron-sized domains in GUVs grown from only a single phospholipid. The membrane heterogeneity is visualized by means of a small fraction (0.1 mol%) of a fluorescent lipid dye. For dipalmitoylphosphatidylcholine GUVs, different types of membrane heterogeneities were detected. First, unexpected formation of micronsized dye-depleted fluid domains was observed upon cooling of GUVs to about 10 K above the lipid main phase transition temperature, TM. Further cooling of the GUVs down to the immediate vicinity of TM renders the domains hexagonal, and in addition, stripe-like dye-enriched structures around the domains are detected. The micron-sized fluid domains in quasi single-component GUVs were observed also when using two other lipids. Whereas the stripe structures are related to the phase transition of the lipid, the dye-excluding domains seem to be caused by traces of impurities present in the sugar solutions. Supplementing the solutions with nm-sized liposomes at millimolar lipid concentration suppresses the formation of the micron-sized domains, presumably by providing competitive binding of the impurities to the liposome membrane in excess. It is likely that such traces of impurities can significantly alter lipid phase diagrams and cause differences among reported ones.