title: Triggering and visualizing the aggregation and fusion of lipid membranes in microfluidic chambers.
abstract: 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 approximately 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.
authors.affiliation: Department of Biomedical Engineering, College of Engineering, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Estes
authors.forename: Santiago R
authors.initials: SR
authors.lastname: Lopez
authors.forename: A Oveta
authors.initials: AO
authors.lastname: Fuller
authors.forename: Michael
authors.initials: M
authors.lastname: Mayer
mesh_terms: D004563:Electrochemistry / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D008051:Lipid Bilayers / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry*; D008561:Membrane Fusion; D008839:Microelectrodes; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D008858:Microscopy, Phase-Contrast / Q000379:methods*; D008968:Molecular Conformation
chemical_list: D008051:Lipid Bilayers; D008081:Liposomes