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The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling
Despite an enormous interest in the role of extracellular vesicles, including exosomes, in cancer and their use as biomarkers for diagnosis, prognosis, drug response and recurrence, there is no consensus on dependable isolation protocols. We provide a comparative evaluation of 4 exosome isolation pr...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Co-Action Publishing
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4169610/ https://www.ncbi.nlm.nih.gov/pubmed/25317274 http://dx.doi.org/10.3402/jev.v3.24858 |
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author | Van Deun, Jan Mestdagh, Pieter Sormunen, Raija Cocquyt, Veronique Vermaelen, Karim Vandesompele, Jo Bracke, Marc De Wever, Olivier Hendrix, An |
author_facet | Van Deun, Jan Mestdagh, Pieter Sormunen, Raija Cocquyt, Veronique Vermaelen, Karim Vandesompele, Jo Bracke, Marc De Wever, Olivier Hendrix, An |
author_sort | Van Deun, Jan |
collection | PubMed |
description | Despite an enormous interest in the role of extracellular vesicles, including exosomes, in cancer and their use as biomarkers for diagnosis, prognosis, drug response and recurrence, there is no consensus on dependable isolation protocols. We provide a comparative evaluation of 4 exosome isolation protocols for their usability, yield and purity, and their impact on downstream omics approaches for biomarker discovery. OptiPrep density gradient centrifugation outperforms ultracentrifugation and ExoQuick and Total Exosome Isolation precipitation in terms of purity, as illustrated by the highest number of CD63-positive nanovesicles, the highest enrichment in exosomal marker proteins and a lack of contaminating proteins such as extracellular Argonaute-2 complexes. The purest exosome fractions reveal a unique mRNA profile enriched for translation, ribosome, mitochondrion and nuclear lumen function. Our results demonstrate that implementation of high purification techniques is a prerequisite to obtain reliable omics data and identify exosome-specific functions and biomarkers. |
format | Online Article Text |
id | pubmed-4169610 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Co-Action Publishing |
record_format | MEDLINE/PubMed |
spelling | pubmed-41696102014-10-14 The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling Van Deun, Jan Mestdagh, Pieter Sormunen, Raija Cocquyt, Veronique Vermaelen, Karim Vandesompele, Jo Bracke, Marc De Wever, Olivier Hendrix, An J Extracell Vesicles Original Research Article Despite an enormous interest in the role of extracellular vesicles, including exosomes, in cancer and their use as biomarkers for diagnosis, prognosis, drug response and recurrence, there is no consensus on dependable isolation protocols. We provide a comparative evaluation of 4 exosome isolation protocols for their usability, yield and purity, and their impact on downstream omics approaches for biomarker discovery. OptiPrep density gradient centrifugation outperforms ultracentrifugation and ExoQuick and Total Exosome Isolation precipitation in terms of purity, as illustrated by the highest number of CD63-positive nanovesicles, the highest enrichment in exosomal marker proteins and a lack of contaminating proteins such as extracellular Argonaute-2 complexes. The purest exosome fractions reveal a unique mRNA profile enriched for translation, ribosome, mitochondrion and nuclear lumen function. Our results demonstrate that implementation of high purification techniques is a prerequisite to obtain reliable omics data and identify exosome-specific functions and biomarkers. Co-Action Publishing 2014-09-18 /pmc/articles/PMC4169610/ /pubmed/25317274 http://dx.doi.org/10.3402/jev.v3.24858 Text en © 2014 Jan Van Deun et al. http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution-Noncommercial 3.0 Unported License, permitting all non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Research Article Van Deun, Jan Mestdagh, Pieter Sormunen, Raija Cocquyt, Veronique Vermaelen, Karim Vandesompele, Jo Bracke, Marc De Wever, Olivier Hendrix, An The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title | The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title_full | The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title_fullStr | The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title_full_unstemmed | The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title_short | The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling |
title_sort | impact of disparate isolation methods for extracellular vesicles on downstream rna profiling |
topic | Original Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4169610/ https://www.ncbi.nlm.nih.gov/pubmed/25317274 http://dx.doi.org/10.3402/jev.v3.24858 |
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