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Triggering and Visualizing the Aggregation and Fusion of Lipid Membranes in Microfluidic Chambers

Daniel J. Estes ^*, Santiago R. Lopez , A. Oveta Fuller   and Michael Mayer ^* 

^* Department of Biomedical Engineering and Department of Chemical Engineering, College of Engineering, and Department of Microbiology and Immunology and Program in Cellular and Molecular Biology, School of Medicine, University of Michigan, Ann Arbor, Michigan 48109

Correspondence: Address reprint requests to Michael Mayer, University of Michigan, Depts. of Biomedical Engineering and Chemical Engineering, Gerstacker Building, Rm. 1107, 2200 Bonisteel Blvd., Ann Arbor, MI 48109-2099. Tel.: 734-763-4609; Fax: 734-763-4371; E-mail: mimayer{at}umich.edu.

We present a method that makes it possible to trigger, observe, and quantify membrane aggregation and fusion of giant liposomes in microfluidic chambers. Using electroformation from spin-coated films of lipids on transparent indium tin oxide electrodes, we formed two-dimensional networks of closely packed, surface-attached giant liposomes. We investigated the effects of fusogenic agents by simply flowing these molecules into the chambers and analyzing the resulting shape changes of more than 100 liposomes in parallel. We used this setup to quantify membrane fusion by several well-studied mechanisms, including fusion triggered by Ca²⁺, polyethylene glycol, and biospecific tethering. Directly observing many liposomes simultaneously proved particularly useful for studying fusion events in the presence of low concentrations of fusogenic agents, when fusion was rare and probabilistic. We applied this microfluidic fusion assay to investigate a novel 30-mer peptide derived from a recently identified human receptor protein, B5, that is important for membrane fusion during the entry of herpes simplex virus into host cells. This peptide triggered fusion of liposomes at an 6 times higher probability than control peptides and caused irreversible interactions between adjacent membranes; it was, however, less fusogenic than Ca²⁺ at comparable concentrations. Closely packed, surface-attached giant liposomes in microfluidic chambers offer a method to observe membrane aggregation and fusion in parallel without requiring the use of micromanipulators. This technique makes it possible to characterize rapidly novel fusogenic agents under well-defined conditions.

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