Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery

Abstract Human plasma is a complex fluid, increasingly used for extracellular vesicle (EV) biomarker studies. Our aim was to find a simple EV-enrichment method for reliable quantification of EVs in plasma to be used as biomarker of disease. Plasma of ten healthy subjects was processed using sediment...

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Autores principales:	Marija Holcar, Jana Ferdin, Simona Sitar, Magda Tušek-Žnidarič, Vita Dolžan, Ana Plemenitaš, Ema Žagar, Metka Lenassi
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f269a5ded5fe4d2588a4a370658df111
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spelling	oai:doaj.org-article:f269a5ded5fe4d2588a4a370658df1112021-12-02T11:42:10ZEnrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery10.1038/s41598-020-78422-y2045-2322https://doaj.org/article/f269a5ded5fe4d2588a4a370658df1112020-12-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-78422-yhttps://doaj.org/toc/2045-2322Abstract Human plasma is a complex fluid, increasingly used for extracellular vesicle (EV) biomarker studies. Our aim was to find a simple EV-enrichment method for reliable quantification of EVs in plasma to be used as biomarker of disease. Plasma of ten healthy subjects was processed using sedimentation rate- (sucrose cushion ultracentrifugation—sUC) and size- (size exclusion chromatography—SEC) based methods. According to nanoparticle tracking analysis (NTA), asymmetrical flow field-flow fractionation coupled to detectors (AF4-UV-MALS), miRNA quantification, transmission electron microscopy and enzyme-linked immunosorbent assay, enrichment of EVs from plasma with sUC method lead to high purity of EVs in the samples. High nanoparticle concentrations after SEC resulted from substantial contamination with lipoproteins and other aggregates of EV-like sizes that importantly affect downstream EV quantification. Additionally, sUC EV-enrichment method linked to quantification with NTA or AF4-UV-MALS is repeatable, as the relative standard deviation of EV size measured in independently processed samples from the same plasma source was 5.4% and 2.1% when analyzed by NTA or AF4-UV-MALS, respectively. In conclusion, the sUC EV-enrichment method is compatible with reliable measurement of concentration and size of EVs from plasma and should in the future be tested on larger cohorts in relation to different diseases. This is one of the first studies using AF4-UV-MALS to quantify EVs in blood plasma, which opens new possible clinical utility for the technique.Marija HolcarJana FerdinSimona SitarMagda Tušek-ŽnidaričVita DolžanAna PlemenitašEma ŽagarMetka LenassiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 10, Iss 1, Pp 1-13 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Marija Holcar Jana Ferdin Simona Sitar Magda Tušek-Žnidarič Vita Dolžan Ana Plemenitaš Ema Žagar Metka Lenassi Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
description	Abstract Human plasma is a complex fluid, increasingly used for extracellular vesicle (EV) biomarker studies. Our aim was to find a simple EV-enrichment method for reliable quantification of EVs in plasma to be used as biomarker of disease. Plasma of ten healthy subjects was processed using sedimentation rate- (sucrose cushion ultracentrifugation—sUC) and size- (size exclusion chromatography—SEC) based methods. According to nanoparticle tracking analysis (NTA), asymmetrical flow field-flow fractionation coupled to detectors (AF4-UV-MALS), miRNA quantification, transmission electron microscopy and enzyme-linked immunosorbent assay, enrichment of EVs from plasma with sUC method lead to high purity of EVs in the samples. High nanoparticle concentrations after SEC resulted from substantial contamination with lipoproteins and other aggregates of EV-like sizes that importantly affect downstream EV quantification. Additionally, sUC EV-enrichment method linked to quantification with NTA or AF4-UV-MALS is repeatable, as the relative standard deviation of EV size measured in independently processed samples from the same plasma source was 5.4% and 2.1% when analyzed by NTA or AF4-UV-MALS, respectively. In conclusion, the sUC EV-enrichment method is compatible with reliable measurement of concentration and size of EVs from plasma and should in the future be tested on larger cohorts in relation to different diseases. This is one of the first studies using AF4-UV-MALS to quantify EVs in blood plasma, which opens new possible clinical utility for the technique.
format	article
author	Marija Holcar Jana Ferdin Simona Sitar Magda Tušek-Žnidarič Vita Dolžan Ana Plemenitaš Ema Žagar Metka Lenassi
author_facet	Marija Holcar Jana Ferdin Simona Sitar Magda Tušek-Žnidarič Vita Dolžan Ana Plemenitaš Ema Žagar Metka Lenassi
author_sort	Marija Holcar
title	Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
title_short	Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
title_full	Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
title_fullStr	Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
title_full_unstemmed	Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
title_sort	enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery
publisher	Nature Portfolio
publishDate	2020
url	https://doaj.org/article/f269a5ded5fe4d2588a4a370658df111
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Enrichment of plasma extracellular vesicles for reliable quantification of their size and concentration for biomarker discovery

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