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dc.contributor.authorAsadi, Jalal
dc.contributor.authorFerguson, Sophie
dc.contributor.authorRaja, Hussain
dc.contributor.authorHacker, Christian
dc.contributor.authorMarius, Phedra
dc.contributor.authorWard, Richard
dc.contributor.authorPliotas, Christos
dc.contributor.authorNaismith, James Henderson
dc.contributor.authorLucocq, John
dc.date.accessioned2017-05-11T09:30:22Z
dc.date.available2017-05-11T09:30:22Z
dc.date.issued2017-08
dc.identifier.citationAsadi , J , Ferguson , S , Raja , H , Hacker , C , Marius , P , Ward , R , Pliotas , C , Naismith , J H & Lucocq , J 2017 , ' Enhanced imaging of lipid rich nanoparticles embedded in methylcellulose films for transmission electron microscopy using mixtures of heavy metals ' Micron , vol. 99 , pp. 40-48 . https://doi.org/10.1016/j.micron.2017.03.019en
dc.identifier.issn0968-4328
dc.identifier.otherPURE: 249642134
dc.identifier.otherPURE UUID: 2108482e-21f7-4964-bb65-16496b101ad0
dc.identifier.otherRIS: urn:0EAA2C4ED6D55B1E9673D85F2CF57DAA
dc.identifier.otherScopus: 85017527213
dc.identifier.otherORCID: /0000-0002-4309-4858/work/34737683
dc.identifier.otherWOS: 000402217600006
dc.identifier.urihttp://hdl.handle.net/10023/10746
dc.descriptionThis work was supported by funds from the St Andrews EM facility and the University of St Andrews. CP is a Royal Society of Edinburgh (RSE) Personal Research Fellow, funded by the Scottish Government. CP is also supported by a Tenovus Scotland Award (T15/41). JML was supported by a Programme grant from the Wellcome Trust (Number 045404).en
dc.description.abstractSynthetic and naturally occurring lipid-rich nanoparticles are of wide ranging importance in biomedicine. They include liposomes, bicelles, nanodiscs, exosomes and virus particles. The quantitative study of these particles requires methods for high-resolution visualization of the whole population. One powerful imaging method is cryo-EM of vitrified samples, but this is technically demanding, requires specialized equipment, provides low contrast and does not reveal all particles present in a population. Another approach is classical negative stain-EM, which is more accessible but is difficult to standardize for larger lipidic structures, which are prone to artifacts of structure collapse and contrast variability. A third method uses embedment in methylcellulose films containing uranyl acetate as a contrasting agent. Methylcellulose embedment has been widely used for contrasting and supporting cryosections but only sporadically for visualizing lipid rich vesicular structures such as endosomes and exosomes. Here we present a simple methylcellulose-based method for routine and comprehensive visualization of synthetic lipid rich nanoparticles preparations, such as liposomes, bicelles and nanodiscs. It combines a novel double-staining mixture of uranyl acetate (UA) and tungsten-based electron stains (namely phosphotungstic acid (PTA) or sodium silicotungstate (STA)) with methylcellulose embedment. While the methylcellulose supports the delicate lipid structures during drying, the addition of PTA or STA to UA provides significant enhancement in lipid structure display and contrast as compared to UA alone. This double staining method should aid routine structural evaluation and quantification of lipid rich nanoparticles structures.
dc.format.extent9
dc.language.isoeng
dc.relation.ispartofMicronen
dc.rights© 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/BY/4.0/)en
dc.subjectBicellesen
dc.subjectPhospholipidsen
dc.subjectMembranesen
dc.subjectLipidsen
dc.subjectMethylcelluloseen
dc.subjectUranyl acetateen
dc.subjectPhosphotungstic aciden
dc.subjectSodium silicotungstateen
dc.subjectNegative stainen
dc.subjectLiposomesen
dc.subjectNanodiscsen
dc.subjectQD Chemistryen
dc.subjectQH301 Biologyen
dc.subjectQR Microbiologyen
dc.subjectNDASen
dc.subject.lccQDen
dc.subject.lccQH301en
dc.subject.lccQRen
dc.titleEnhanced imaging of lipid rich nanoparticles embedded in methylcellulose films for transmission electron microscopy using mixtures of heavy metalsen
dc.typeJournal articleen
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews.School of Medicineen
dc.contributor.institutionUniversity of St Andrews.Biomedical Sciences Research Complexen
dc.contributor.institutionUniversity of St Andrews.School of Biologyen
dc.contributor.institutionUniversity of St Andrews.School of Chemistryen
dc.contributor.institutionUniversity of St Andrews.EaSTCHEMen
dc.contributor.institutionUniversity of St Andrews.Cellular Medicine Divisionen
dc.identifier.doihttps://doi.org/10.1016/j.micron.2017.03.019
dc.description.statusPeer revieweden
dc.identifier.urlhttp://europepmc.org/articles/PMC5465805en


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