University of Oulu

Ullah, M. S., Zhivonitko, V. V., Samoylenko, A., Zhyvolozhnyi, A., Viitala, S., Kankaanpää, S., Komulainen, S., Schröder, L., Vainio, S. J., & Telkki, V.-V. (2021). Identification of extracellular nanoparticle subsets by nuclear magnetic resonance. Chemical Science, 12(24), 8311–8319.

Identification of extracellular nanoparticle subsets by nuclear magnetic resonance

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Author: Ullah, Md Sharif1; Zhivonitko, Vladimir V.1; Samoylenko, Anatoliy2;
Organizations: 1NMR Research Unit, University of Oulu, Finland
2Laboratory of Developmental Biology, Infotech Oulu, Oulu Center for Cell-Matrix Research, Kvantum Institute, Faculty of Biochemistry and Molecular Medicine, Oulu, Finland
3Production Systems, Natural Resources Institute Finland (Luke), Jokioinen, Finland
4Molecular Imaging, Leibniz-Forschungsinstitut f¨ur Molekulare Pharmakologie (FMP), Berlin, Germany
5Division of Translational Molecular Imaging, German Cancer Research Center (DKFZ), Heidelberg, Germany
Format: article
Version: published version
Access: open
Online Access: PDF Full Text (PDF, 1.3 MB)
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Language: English
Published: Royal Society of Chemistry, 2021
Publish Date: 2021-06-29


Exosomes are a subset of secreted lipid envelope-encapsulated extracellular vesicles (EVs) of 50–150 nm diameter that can transfer cargo from donor to acceptor cells. In the current purification protocols of exosomes, many smaller and larger nanoparticles such as lipoproteins, exomers and microvesicles are typically co-isolated as well. Particle size distribution is one important characteristics of EV samples, as it reflects the cellular origin of EVs and the purity of the isolation. However, most of the physicochemical analytical methods today cannot illustrate the smallest exosomes and other small particles like the exomers. Here, we demonstrate that diffusion ordered spectroscopy (DOSY) nuclear magnetic resonance (NMR) method enables the determination of a very broad distribution of extracellular nanoparticles, ranging from 1 to 500 nm. The range covers sizes of all particles included in EV samples after isolation. The method is non-invasive, as it does not require any labelling or other chemical modification. We investigated EVs secreted from milk as well as embryonic kidney and renal carcinoma cells. Western blot analysis and immuno-electron microscopy confirmed expression of exosomal markers such as ALIX, TSG101, CD81, CD9, and CD63 in the EV samples. In addition to the larger particles observed by nanoparticle tracking analysis (NTA) in the range of 70–500 nm, the DOSY distributions include a significant number of smaller particles in the range of 10–70 nm, which are visible also in transmission electron microscopy images but invisible in NTA. Furthermore, we demonstrate that hyperpolarized chemical exchange saturation transfer (Hyper-CEST) with ¹²⁹Xe NMR indicates also the existence of smaller and larger nanoparticles in the EV samples, providing also additional support for DOSY results. The method implies also that the Xe exchange is significantly faster in the EV pool than in the lipoprotein/exomer pool.

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Series: Chemical science
ISSN: 2041-6520
ISSN-E: 2041-6539
ISSN-L: 2041-6520
Volume: 12
Issue: 24
Pages: 8311 - 8319
DOI: 10.1039/D1SC01402A
Type of Publication: A1 Journal article – refereed
Field of Science: 114 Physical sciences
116 Chemical sciences
Funding: We acknowledge the financial support of the European Research Council (ERC) under Horizon 2020 (H2020/2018–2022/ERC grant agreement no. 772110), Academy of Finland (grants #289649, 294027, 319216 and 323480), EU FET Open grant 829040, Sigrid Juselius Foundation, Cancer Foundation Finland (grants 2017 and 2018), the Kvantum institute (University of Oulu), Ministry of Agriculture and Forestry of Finland, the Deutsche Forschungsgemeinschaft (Koselleck grant no. SCHR 995/5-1), and the Dieter Morszeck Stiftung.
EU Grant Number: (772110) UFLNMR - Ultrafast Laplace NMR
(829040) MindGAP - Bridging the gap between Mind, Brain and Body: exosome role and monitoring
Academy of Finland Grant Number: 289649
Detailed Information: 289649 (Academy of Finland Funding decision)
294027 (Academy of Finland Funding decision)
319216 (Academy of Finland Funding decision)
323480 (Academy of Finland Funding decision)
Copyright information: © 2021 The Author(s). Published by the Royal Society of Chemistry. This article is licensed under a Creative Commons Attribution 3.0 Unported Licence.