Abstract
Liposomes are versatile supramolecular assemblies widely used in basic and applied sciences. Here we present a novel microfluidics-based method, octanol-assisted liposome assembly (OLA), to form monodisperse, cell-sized (5–20 μm), unilamellar liposomes with excellent encapsulation efficiency. Akin to bubble blowing, an inner aqueous phase and a surrounding lipid-carrying 1-octanol phase is pinched off by outer fluid streams. Such hydrodynamic flow focusing results in double-emulsion droplets that spontaneously develop a side-connected 1-octanol pocket. Owing to interfacial energy minimization, the pocket splits off to yield fully assembled solvent-free liposomes within minutes. This solves the long-standing fundamental problem of prolonged presence of residual oil in the liposome bilayer. We demonstrate the unilamellarity of liposomes with functional α-haemolysin protein pores in the membrane and validate the biocompatibility by inner leaflet localization of bacterial divisome proteins (FtsZ and ZipA). OLA offers a versatile platform for future analytical tools, delivery systems, nanoreactors and synthetic cells.
Highlights
Liposomes are versatile supramolecular assemblies widely used in basic and applied sciences
Double-emulsion droplets formed in octanol-assisted liposome assembly (OLA) quickly develop into an intermediate complex of an aqueous lumen encapsulated by a lipid bilayer interface and a 1-octanol pocket connected to it
We demonstrate the localization of encapsulated bacterial divisome proteins (FtsZ and ZipA) to the membrane, which serves to show the biocompatibility of the vesicles
Summary
Formed a prominent side pocket, the volume of the dispensed LO phase was smaller in case of Fig. 3a, and the pocket developed afterwards, not within the displayed time range. Note that producing small liposomes in a controlled manner is highly important when working with protein systems that naturally function at this range of cell sizes and membrane curvature. We tested the biocompatibility of OLA-based liposomes by encapsulating two key bacterial divisome proteins, viz., FtsZ (labelled with Alexa Fluor 488) and sZipA (labelled with Alexa Fluor 647), inside the liposomes. When DGS-NTA(Ni) lipids were absent from the bilayer, FtsZ filaments and bundles were randomly distributed inside the liposomes (Fig. 6b). These experiments clearly show that OLA-based liposomes can efficiently encapsulate active proteins and present a functional lipid bilayer suitable for interactions with biomolecules. The mixture of DOPC and DGS-NTA(Ni) worked very well (Fig. 6)
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