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Management of the diffusion of 4-methylumbelliferone across phases in microdroplet-based systems for in vitro protein evolution

Wu, Nan; Courtois, Fabienne; Zhu, Yonggang; Oakeshott, John Graham; Easton, Christopher; Abell, Christopher

Description

Fluorongenic reagents based on 4-methylumbelliferone (4-MU) have been widely used for the detection of phosphatase, sulfatase, esterase, lipase and glycosidase activities in conventionally formatted enzyme assay systems. However, the sensitivity of assays based on these substrates is also potentially very useful in the microdroplet formats now being developed for high throughput in vitro evolution experiments. In this article, we report the investigation of diffusion of 4-MU as a model dye from...[Show more]

dc.contributor.authorWu, Nan
dc.contributor.authorCourtois, Fabienne
dc.contributor.authorZhu, Yonggang
dc.contributor.authorOakeshott, John Graham
dc.contributor.authorEaston, Christopher
dc.contributor.authorAbell, Christopher
dc.date.accessioned2015-12-10T22:41:09Z
dc.identifier.issn0173-0835
dc.identifier.urihttp://hdl.handle.net/1885/57761
dc.description.abstractFluorongenic reagents based on 4-methylumbelliferone (4-MU) have been widely used for the detection of phosphatase, sulfatase, esterase, lipase and glycosidase activities in conventionally formatted enzyme assay systems. However, the sensitivity of assays based on these substrates is also potentially very useful in the microdroplet formats now being developed for high throughput in vitro evolution experiments. In this article, we report the investigation of diffusion of 4-MU as a model dye from water-in-oil droplets and the internal aqueous phase of water-in-oil-in-water droplets in microfluidics. The effect of BSA in the aqueous phase on the diffusion of 4-MU is also discussed. Based on these results, we provided here proof-of-concept of the reaction of the enzyme OpdA with the substrate coumaphos in water-in-oil-in-water droplets. In this double-emulsion system, the reaction of OpdA and coumaphos was achieved by allowing coumaphos to diffuse from the continuous aqueous phase across the oil phase into the internal aqueous droplets.
dc.publisherWiley-VCH Verlag GMBH
dc.sourceElectrophoresis
dc.subjectKeywords: coumafos; hymecromone; phosphotriesterase; water oil cream; article; diffusion; enzyme substrate complex; microfluidic analysis; phase transition; Animals; Cattle; Coumaphos; Diffusion; Emulsions; Enzyme Assays; Fluorescent Dyes; Hymecromone; Micelles; Mi Directed evolution; Microfluidics; Single- and double-emulsion droplets; Substrate diffusion
dc.titleManagement of the diffusion of 4-methylumbelliferone across phases in microdroplet-based systems for in vitro protein evolution
dc.typeJournal article
local.description.notesImported from ARIES
local.identifier.citationvolume31
dc.date.issued2010
local.identifier.absfor030302 - Nanochemistry and Supramolecular Chemistry
local.identifier.ariespublicationu4005981xPUB414
local.type.statusPublished Version
local.contributor.affiliationWu, Nan, College of Physical and Mathematical Sciences, ANU
local.contributor.affiliationCourtois, Fabienne, University of Cambridge
local.contributor.affiliationZhu, Yonggang, CSIRO Materials Science and Engineering
local.contributor.affiliationOakeshott, John Graham, CSIRO Division of Entomology
local.contributor.affiliationEaston, Christopher, College of Physical and Mathematical Sciences, ANU
local.contributor.affiliationAbell, Christopher, University of Cambridge
local.description.embargo2037-12-31
local.bibliographicCitation.issue18
local.bibliographicCitation.startpage3121
local.bibliographicCitation.lastpage3128
local.identifier.doi10.1002/elps.201000140
local.identifier.absseo970103 - Expanding Knowledge in the Chemical Sciences
dc.date.updated2016-02-24T10:23:40Z
local.identifier.scopusID2-s2.0-77956940537
local.identifier.thomsonID000283389900015
CollectionsANU Research Publications

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