title: Nerve cell-mimicking liposomes as biosensor for botulinum neurotoxin complete physiological activity.
abstract: Botulinum neurotoxins (BoNT) are the most toxic substances known, and their neurotoxic properties and paralysing effects are exploited for medical treatment of a wide spectrum of disorders. To accurately quantify the potency of a pharmaceutical BoNT preparation, its physiological key activities (binding to membrane receptor, translocation, and proteolytic degradation of SNARE proteins) need to be determined. To date, this was only possible using animal models, or, to a limited extent, cell-based assays. We here report a novel in vitro system for BoNT/B analysis, based on nerve-cell mimicking liposomes presenting motoneuronal membrane receptors required for BoNT binding. Following triggered membrane translocation of the toxin's Light Chain, the endopeptidase activity can be quantitatively monitored employing a FRET-based reporter assay within the functionalized liposomes. We were able to detect BoNT/B physiological activity at picomolar concentrations in short time, opening the possibility for future replacement of animal experimentation in pharmaceutical BoNT testing.
authors.affiliation: Institute for Food, Nutrition and Health, ETH Zurich, Schmelzbergstrasse 7, CH-8092 Zurich, Switzerland. Electronic address: Oliver.Weingart@hest.ethz.ch.
authors.forename: Oliver G
authors.initials: OG
authors.lastname: Weingart
authors.affiliation: Institute for Food, Nutrition and Health, ETH Zurich, Schmelzbergstrasse 7, CH-8092 Zurich, Switzerland.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Loessner
mesh_terms: D000826:Animal Testing Alternatives; D015374:Biosensing Techniques; D001905:Botulinum Toxins / Q000633:toxicity*; D010450:Endopeptidases / Q000378:metabolism; D008081:Liposomes; D008855:Microscopy, Electron, Scanning; D009474:Neurons / Q000187:drug effects*; D009498:Neurotoxins / Q000633:toxicity*; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D008081:Liposomes; D009498:Neurotoxins; D010450:Endopeptidases; D001905:Botulinum Toxins
keywords: Animal testing alternatives; Botulinum neurotoxin; In vitro; Liposome; Potency testing