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Communication

The Separation and Characterization of Extracellular Vesicles from Medium Conditioned by Bovine Embryos

1
Department of Reproduction, Obstetrics and Herd Health, Faculty of Veterinary Medicine, University of Ghent, Salisburylaan 133, B-9820 Merelbeke, Belgium
2
Department of Nutrition, Genetics and Ethology, Faculty of Veterinary Medicine, Ghent University, B-9820 Merelbeke, Belgium
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Plant Diseases and Crop Protection, Institute of Crop Science and Resource Conservation, University of Bonn, Nussallee 9, 53115 Bonn, Germany
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Department of Morphology-Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke, Belgium
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Laboratory of Experimental Cancer Research, Department of Human Structure and Repair, Ghent University, B-9000 Ghent, Belgium
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Cancer Research Institute Ghent (CRIG), B-9000 Ghent, Belgium
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2020, 21(8), 2942; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21082942
Received: 31 March 2020 / Revised: 17 April 2020 / Accepted: 21 April 2020 / Published: 22 April 2020
(This article belongs to the Special Issue Extracellular Vesicles in Reproduction)
Extracellular vesicles (EVs) have been identified as one of the communication mechanisms amongst embryos. They are secreted into the embryo culture medium and, as such, represent a source of novel biomarkers for identifying the quality of cells and embryos. However, only small amounts of embryo-conditioned medium are available, which represents a challenge for EV enrichment. Our aim is to assess the suitability of different EV separation methods to retrieve EVs with high specificity and sufficient efficiency. Bovine embryo-conditioned medium was subjected to differential ultracentrifugation (DU), OptiPrepTM density gradient (ODG) centrifugation, and size exclusion chromatography. Separated EVs were characterized by complementary characterization methods, including Western blot, electron microscopy, and nanoparticle tracking analysis, to assess the efficiency and specificity. OptiPrepTM density gradient centrifugation outperformed DU and SEC in terms of specificity by substantial removal of contaminating proteins such as ribonucleoprotein complexes (Argonaute-2 (AGO-2)) and lipoproteins (ApoA-I) from bovine embryo-derived EVs (density: 1.02–1.04, 1.20–1.23 g/mL, respectively). In conclusion, ODG centrifugation is the preferred method for identifying EV-enriched components and for improving our understanding of EV function in embryo quality and development. View Full-Text
Keywords: extracellular vesicles; isolation; protocols; ultracentrifugation; size exclusion chromatography; OptiprepTM density gradient extracellular vesicles; isolation; protocols; ultracentrifugation; size exclusion chromatography; OptiprepTM density gradient
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MDPI and ACS Style

Pavani, K.C.; Lin, X.; Hamacher, J.; Broeck, W.V.D.; Couck, L.; Peelman, L.; Hendrix, A.; Van Soom, A. The Separation and Characterization of Extracellular Vesicles from Medium Conditioned by Bovine Embryos. Int. J. Mol. Sci. 2020, 21, 2942. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21082942

AMA Style

Pavani KC, Lin X, Hamacher J, Broeck WVD, Couck L, Peelman L, Hendrix A, Van Soom A. The Separation and Characterization of Extracellular Vesicles from Medium Conditioned by Bovine Embryos. International Journal of Molecular Sciences. 2020; 21(8):2942. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21082942

Chicago/Turabian Style

Pavani, Krishna C., Xiaoyuan Lin, Joachim Hamacher, Wim V.D. Broeck, Liesbeth Couck, Luc Peelman, An Hendrix, and Ann Van Soom. 2020. "The Separation and Characterization of Extracellular Vesicles from Medium Conditioned by Bovine Embryos" International Journal of Molecular Sciences 21, no. 8: 2942. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21082942

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