A Novel and Versatile Approach for Preparation of Single-Vesicle Arrays

Abstract

Giant unilamellar vesicles (GUVs) are attractive model membranes, commonly used to study membrane processes such as membrane fusion and protein-lipid interactions. Arrays of GUVs are, therefore, ideal platforms for high-throughput screening of such processes. Here, we present a simple approach to form arrays of GUVs with or without membrane-proteins. This approach combines hydrogel-based microcontact printing with the commonly-used electroformation technique to generate arrays of GUVs with controllable size and composition. First, micro-patterned agarose stamps deliver aqueous solutions of small liposomes/proteoliposomes onto conductive substrates. GUVs are then produced from these patterned deposits using electroformation. By varying the feature size of stamps, we controlled the number of liposomes grown per deposit and ultimately formed single-vesicle arrays with a narrow size distribution (37±1μm diameter). We demonstrated this method's capability to create giant proteoliposome arrays by electroforming vesicles from stamped aquaporin-containing membrane deposits, and acetylcholine receptor-containing cell membrane fragments and confirmed the presence of these proteins by immunofluorescent-assays. We further illustrated the versatility of this approach by forming GUV arrays in physiologically-relevant solutions. To demonstrate this system's applicability for studying protein-lipid interactions, we investigated the interactions of a membrane-associated protein, annexin-V, with phosphatidylserine-containing GUVs.View Large Image | View Hi-Res Image | Download PowerPoint Slide