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Separation of small extracellular vesicles (sEV) from human blood by Superose 6 size exclusion chromatography

Authors :
Jerome Nouvel
Gonzalo Bustos Quevedo
Tony Prinz
Ramsha Masood
George Daaboul
Tanja Gainey‐Schleicher
Uwe Wittel
Sophia Chikhladze
Bence Melykuti
Martin Helmstaedter
Karl Winkler
Irina Nazarenko
Gerhard Pütz
Source :
Journal of Extracellular Vesicles, Vol 13, Iss 10, Pp n/a-n/a (2024)
Publication Year :
2024
Publisher :
Wiley, 2024.

Abstract

Abstract Extracellular vesicles (EVs) are valuable targets for liquid biopsy. However, attempts to introduce EV‐based biomarkers into clinical practice have not been successful to the extent expected. One of the reasons for this failure is the lack of reliable methods for EV baseline purification from complex biofluids, such as cell‐free plasma or serum. Because available one‐step approaches for EV isolation are insufficient to purify EVs, the majority of studies on clinical samples were performed either on a mixture of EVs and lipoproteins, whilst the real number of EVs and their individual specific biomarker content remained elusive, or on a low number of samples of sufficient volume to allow elaborate 2‐step EV separation by size and density, resulting in a high purity but utmost low recovery. Here we introduce Fast Protein Liquid Chromatography (FPLC) using Superose 6 as a matrix to obtain small EVs from biofluids that are almost free of soluble proteins and lipoproteins. Along with the estimation of a realistic number of small EVs in human samples, we show temporal resolution of the effect of the duration of postprandial phase on the proportion of lipoproteins in purified EVs, suggesting acceptable time frames additionally to the recommendation to use fasting samples for human studies. Furthermore, we assessed a potential value of pure EVs for liquid biopsy, exemplarily examining EV‐ and tumour‐biomarkers in pure FPLC‐derived fractions isolated from the serum of patients with pancreatic cancer. Consistent among different techniques, showed the presence of diseases‐associated biomarkers in pure EVs, supporting the feasibility of using single‐vesicle analysis for liquid biopsy.

Details

Language :
English
ISSN :
20013078
Volume :
13
Issue :
10
Database :
Directory of Open Access Journals
Journal :
Journal of Extracellular Vesicles
Publication Type :
Academic Journal
Accession number :
edsdoj.588507e5d9a54be8a0d1eefab47b6573
Document Type :
article
Full Text :
https://doi.org/10.1002/jev2.70008