Recombinant extracellular vesicles as biological reference material for method development, data normalization and assessment of (pre-)analytical variables.
Journal
Nature protocols
ISSN: 1750-2799
Titre abrégé: Nat Protoc
Pays: England
ID NLM: 101284307
Informations de publication
Date de publication:
02 2021
02 2021
Historique:
received:
27
02
2020
accepted:
15
10
2020
pubmed:
17
1
2021
medline:
9
3
2021
entrez:
16
1
2021
Statut:
ppublish
Résumé
The diagnostic and therapeutic use of extracellular vesicles (EV) is under intense investigation and may lead to societal benefits. Reference materials are an invaluable resource for developing, improving and assessing the performance of regulated EV applications and for quantitative and objective data interpretation. We have engineered recombinant EV (rEV) as a biological reference material. rEV have similar biochemical and biophysical characteristics to sample EV and function as an internal quantitative and qualitative control throughout analysis. Spiking rEV in bodily fluids prior to EV analysis maps technical variability of EV applications and promotes intra- and inter-laboratory studies. This protocol, which is an Extension to our previously published protocol (Tulkens et al., 2020), describes the production, separation and quality assurance of rEV, their dilution and addition to bodily fluids, and the detection steps based on complementary fluorescence, nucleic acid and protein measurements. We demonstrate the use of rEV for method development, data normalization and assessment of pre-analytical variables. The protocol can be adopted by researchers with standard laboratory and basic EV separation/characterization experience and requires ~4-5 d.
Identifiants
pubmed: 33452501
doi: 10.1038/s41596-020-00446-5
pii: 10.1038/s41596-020-00446-5
doi:
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
603-633Références
van Niel, G., D’Angelo, G. & Raposo, G. Shedding light on the cell biology of extracellular vesicles. Nat. Rev. Mol. Cell Biol. 19, 213–228 (2018).
pubmed: 29339798
doi: 10.1038/nrm.2017.125
Kalluri, R. & LeBleu, V. S. The biology, function, and biomedical applications of exosomes. Science 367, eaau6977 (2020).
Melo, S. A. et al. Glypican-1 identifies cancer exosomes and detects early pancreatic cancer. Nature 523, 177–182 (2015).
pubmed: 26106858
pmcid: 4825698
doi: 10.1038/nature14581
Hoshino, A. et al. Tumour exosome integrins determine organotropic metastasis. Nature 527, 329–335 (2015).
pubmed: 26524530
pmcid: 4788391
doi: 10.1038/nature15756
Peinado, H. et al. Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET. Nat. Med. 18, 883–891 (2012).
pubmed: 22635005
pmcid: 3645291
doi: 10.1038/nm.2753
Kamerkar, S. et al. Exosomes facilitate therapeutic targeting of oncogenic KRAS in pancreatic cancer. Nature 546, 498–503 (2017).
pubmed: 28607485
pmcid: 5538883
doi: 10.1038/nature22341
Hill, A. F. Extracellular vesicles and neurodegenerative diseases. J. Neurosci. 39, 9269–9273 (2019).
pubmed: 31748282
pmcid: 6867808
doi: 10.1523/JNEUROSCI.0147-18.2019
Cosenza, S. et al. Mesenchymal stem cells-derived exosomes are more immunosuppressive than microparticles in inflammatory arthritis. Theranostics 8, 1399–1410 (2018).
pubmed: 29507629
pmcid: 5835945
doi: 10.7150/thno.21072
Hergenreider, E. et al. Atheroprotective communication between endothelial cells and smooth muscle cells through miRNAs. Nat. Cell Biol. 14, 249–256 (2012).
pubmed: 22327366
doi: 10.1038/ncb2441
Mendt, M. et al. Generation and testing of clinical-grade exosomes for pancreatic cancer. JCI Insight 3, e99263 (2018).
pmcid: 5931131
doi: 10.1172/jci.insight.99263
De Wever, O. & Hendrix, A. A supporting ecosystem to mature extracellular vesicles into clinical application. EMBO J. 38, e101412 (2019).
Van Deun, J. et al. The impact of disparate isolation methods for extracellular vesicles on downstream RNA Profiling. J. Extracell. Vesicles 3, 24858 (2014).
doi: 10.3402/jev.v3.24858
Kowal, J. et al. Proteomic comparison defines novel markers to characterize heterogeneous populations of extracellular vesicle subtypes. Proc. Natl Acad. USA 113, 968–977 (2016).
doi: 10.1073/pnas.1521230113
Van Deun, J. et al. EV-TRACK: transparent reporting and centralizing knowledge in extracellular vesicle research. Nat. Methods 14, 228–232 (2017).
pubmed: 28245209
doi: 10.1038/nmeth.4185
Simonsen, J. B. What are we looking at? Extracellular vesicles, lipoproteins, or both? Circ. Res. 121, 920–922 (2017).
pubmed: 28963190
doi: 10.1161/CIRCRESAHA.117.311767
Jeppesen, D. K. et al. Reassessment of exosome composition. Cell 177, 428–445.e18 (2019).
pubmed: 30951670
pmcid: 6664447
doi: 10.1016/j.cell.2019.02.029
Tulkens, J. et al. Increased levels of systemic LPS-positive bacterial extracellular vesicles in patients with intestinal barrier dysfunction. Gut 69, 191–193 (2018).
pubmed: 30518529
pmcid: 6943244
doi: 10.1136/gutjnl-2018-317726
Geeurickx, E. & Hendrix, A. Targets, pitfalls and reference materials for liquid biopsy tests in cancer diagnostics. Mol. Aspects Med. 72, 100828 (2019).
Valkonen, S. et al. Biological reference materials for extracellular vesicle studies. Eur. J. Pharm. Sci. 98, 4–16 (2017).
pubmed: 27622921
doi: 10.1016/j.ejps.2016.09.008
Geeurickx, E. et al. The generation and use of recombinant extracellular vesicles as biological reference material. Nat. Commun. 10, 3288 (2019).
pubmed: 31337761
pmcid: 6650486
doi: 10.1038/s41467-019-11182-0
van der Pol, E., Coumans, F. A. W., Sturk, A., Nieuwland, R. & Van Leeuwen, T. G. Refractive index determination of nanoparticles in suspension using nanoparticle tracking analysis. Nano Lett. 14, 6195–6201 (2014).
pubmed: 25256919
doi: 10.1021/nl503371p
Gardiner, C. et al. Measurement of refractive index by nanoparticle tracking analysis reveals heterogeneity in extracellular vesicles. J. Extracell. Vesicles 3, 25361 (2014).
pubmed: 25425324
doi: 10.3402/jev.v3.25361
Tulkens, J., De Wever, O. & Hendrix, A. Analyzing bacterial extracellular vesicles in human body fluids by orthogonal biophysical separation and biochemical characterization. Nat. Protoc. 15, 40–67 (2020).
pubmed: 31776460
doi: 10.1038/s41596-019-0236-5
Varga, Z. et al. Hollow organosilica beads as reference particles for optical detection of extracellular vesicles. J. Thromb. Haemost. 16, 1646–1655 (2018).
doi: 10.1111/jth.14193
Lapinski, M. M., Castro-Forero, A., Greiner, A. J., Ofoli, R. Y. & Blanchard, G. J. Comparison of liposomes formed by sonication and extrusion: rotational and translational diffusion of an embedded chromophore. Langmuir 23, 11677–11678 (2007).
pubmed: 17939695
doi: 10.1021/la7020963
Lozano-Andrés, E. et al. Tetraspanin-decorated extracellular vesicle-mimetics as a novel adaptable reference material. J. Extracell. Vesicles 8, 1573052 (2019).
pubmed: 30863514
pmcid: 6407598
doi: 10.1080/20013078.2019.1573052
Lane, R. E., Korbie, D., Anderson, W., Vaidyanathan, R. & Trau, M. Analysis of exosome purification methods using a model liposome system and tunable-resistive pulse sensing. Sci. Rep. 5, 7639 (2015).
pubmed: 25559219
pmcid: 4648344
doi: 10.1038/srep07639
Görgens, A. et al. Optimisation of imaging flow cytometry for the analysis of single extracellular vesicles by using fluorescence-tagged vesicles as biological reference material. J. Extracell. Vesicles 8, 1587567 (2019).
pubmed: 30949308
pmcid: 6442110
doi: 10.1080/20013078.2019.1587567
Lai, C. P. et al. Visualization and tracking of tumour extracellular vesicle delivery and RNA translation using multiplexed reporters. Nat. Commun. 6, 7029 (2015).
pubmed: 25967391
pmcid: 4435621
doi: 10.1038/ncomms8029
van der Vlist, E. J., Nolte-’t Hoen, E. N. M., Stoorvogel, W., Arkesteijn, G. J. A. & Wauben, M. H. M. Fluorescent labeling of nano-sized vesicles released by cells and subsequent quantitative and qualitative analysis by high-resolution flow cytometry. Nat. Protoc. 7, 1311–1326 (2012).
pubmed: 22722367
doi: 10.1038/nprot.2012.065
Tang, V. A. et al. Engineered retroviruses as fluorescent biological reference particles for nanoscale flow cytometry. Preprint at bioRxiv https://doi.org/10.1101/614461 (2019).
Gould, S. J., Booth, A. M. & Hildreth, J. E. K. The Trojan exosome hypothesis. Proc. Natl Acad. Sci. USA 100, 10592–10597 (2003).
pubmed: 12947040
doi: 10.1073/pnas.1831413100
Fujii, K., Hurley, J. H. & Freed, E. O. Beyond Tsg101: the role of Alix in ‘ESCRTing’ HIV-1. Nat. Rev. Microbiol. 5, 912–916 (2007).
pubmed: 17982468
doi: 10.1038/nrmicro1790
de Rond, L. et al. Comparison of generic fluorescent markers for detection of extracellular vesicles by flow cytometry. Clin. Chem. 64, 680–689 (2018).
pubmed: 29453194
doi: 10.1373/clinchem.2017.278978
Théry, C. et al. Minimal information for studies of extracellular vesicles 2018 (MISEV2018): a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines. J. Extracell. Vesicles 7, 1535750 (2018).
pubmed: 30637094
pmcid: 6322352
doi: 10.1080/20013078.2018.1535750
Coumans, F. A. W. et al. Methodological guidelines to study extracellular vesicles. Circ. Res. 120, 1632–1648 (2017).
pubmed: 28495994
doi: 10.1161/CIRCRESAHA.117.309417
Dettenhofer, M. & Yu, X. F. Highly purified human immunodeficiency virus type 1 reveals a virtual absence of Vif in virions. J. Virol. 73, 1460–1467 (1999).
pubmed: 9882352
pmcid: 103971
doi: 10.1128/JVI.73.2.1460-1467.1999
Jeyaram, A. & Jay, S. M. Preservation and storage stability of extracellular vesicles for therapeutic applications. AAPS J. 20, 1 (2018).
doi: 10.1208/s12248-017-0160-y
Lorincz, Á. M. et al. Effect of storage on physical and functional properties of extracellular vesicles derived from neutrophilic granulocytes. J. Extracell. Vesicles 3, 25465 (2014).
pubmed: 25536933
doi: 10.3402/jev.v3.25465
Dhondt, B. et al. Unravelling the proteomic landscape of extracellular vesicles in prostate cancer by density-based fractionation of urine. J. Extracell. Vesicles 9, 1736935 (2020).
pubmed: 32284825
pmcid: 7144211
doi: 10.1080/20013078.2020.1736935
Suk, J. S., Xu, Q., Kim, N., Hanes, J. & Ensign, L. M. PEGylation as a strategy for improving nanoparticle-based drug and gene delivery. Adv. Drug Deliv. Rev. 99, 28–51 (2016).
pubmed: 26456916
doi: 10.1016/j.addr.2015.09.012
van der Pol, E. et al. Particle size distribution of exosomes and microvesicles determined by transmission electron microscopy, flow cytometry, nanoparticle tracking analysis, and resistive pulse sensing. J. Thromb. Haemost. 12, 1182–1192 (2014).
pubmed: 24818656
doi: 10.1111/jth.12602
Zhu, J. Mammalian cell protein expression for biopharmaceutical production. Biotechnol. Adv. 30, 1158–1170 (2012).
pubmed: 21968146
doi: 10.1016/j.biotechadv.2011.08.022
Lin, Y. C. et al. Genome dynamics of the human embryonic kidney 293 lineage in response to cell biology manipulations. Nat. Commun. 5, 4767 (2014).
pubmed: 25182477
pmcid: 4166678
doi: 10.1038/ncomms5767
Kräusslich, H. G. et al. Analysis of protein expression and virus-like particle formation in mammalian cell lines stably expressing HIV-1 gag and env gene products with or without active HIV proteinase. Virology 192, 605–617 (1993).
pubmed: 8421902
doi: 10.1006/viro.1993.1077
Nie, Z. et al. HIV-1 protease processes procaspase 8 to cause mitochondrial release of cytochrome c, caspase cleavage and nuclear fragmentation. Cell Death Differ. 9, 1172–1184 (2002).
pubmed: 12404116
doi: 10.1038/sj.cdd.4401094
Titeca, K. et al. Analyzing trapped protein complexes by Virotrap and SFINX. Nat. Protoc. 12, 881–898 (2017).
pubmed: 28358392
doi: 10.1038/nprot.2017.014
Young, A. T. L., Moore, R. B., Murray, A. G., Mullen, J. C. & Lakey, J. R. T. Assessment of different transfection parameters in efficiency optimization. Cell Transplant. 13, 179–185 (2004).
pubmed: 15129764
doi: 10.3727/000000004773301861
Vergauwen, G. et al. Confounding factors of ultrafiltration and protein analysis in extracellular vesicle research. Sci. Rep. 7, 2704 (2017).
pubmed: 28577337
pmcid: 5457435
doi: 10.1038/s41598-017-02599-y
Livshts, M. A. et al. Isolation of exosomes by differential centrifugation: theoretical analysis of a commonly used protocol. Sci. Rep. 5, 17319 (2015).
pubmed: 26616523
pmcid: 4663484
doi: 10.1038/srep17319
Maas, S. L. N. et al. Possibilities and limitations of current technologies for quantification of biological extracellular vesicles and synthetic mimics. J. Control. Release 200, 87–96 (2015).
pubmed: 25555362
pmcid: 4324667
doi: 10.1016/j.jconrel.2014.12.041
Bustin, S. A. et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin. Chem. 55, 611–622 (2009).
pubmed: 19246619
doi: 10.1373/clinchem.2008.112797
Quah, B. J. C. & O’Neill, H. C. Mycoplasma contaminants present in exosome preparations induce polyclonal B cell responses. J. Leukoc. Biol. 82, 1070–1082 (2007).
pubmed: 17698916
doi: 10.1189/jlb.0507277
Mahmood, T. & Yang, P.-C. Western blot: technique, theory, and trouble shooting. N. Am. J. Med. Sci. 4, 429–434 (2012).