Abstract
Extracellular vesicles (EVs), including exosomes, are nano-sized membrane-bound particles which are released by cells. They have been found in all examined body fluids and can be isolated from conditioned cell culture media. These vesicles have gained increasing attention due to their importance in cellular cross talk, in both health and disease. For example, keratinocyte-derived EVs have been described to modulate melanin production in epidermis. Similar EVs were also shown to have an important role in skin immunology, by stimulating dendritic cells. In this chapter, we will describe how to isolate EVs from keratinocyte cultures and how to perform characterization by Western blot, nanoparticle tracking analysis, and transmission electron microscopy.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Change history
11 December 2019
The original version of this chapter was inadvertently published with incorrect spelling of surname of the authors. The names should read Sebastian Sjöqvist, Aya Imafuku, Danu Gupta, and Samir EL Andaloussi, and not Sebastian Sjöqvist, Aya Imafuku, Dhanu Ghupta, and Samir E. L. Andaloussi.
References
Wiklander OPB, Brennan MÁ, Lötvall J et al (2019) Advances in therapeutic applications of extracellular vesicles. Sci Transl Med 11:eaav8521
Andaloussi SEL, Mäger I, Breakefield XO et al (2013) Extracellular vesicles: biology and emerging therapeutic opportunities. Nat Rev Drug Discov 12:347–357
Otake K, Kamiguchi H, Hirozane Y (2019) Identification of biomarkers for amyotrophic lateral sclerosis by comprehensive analysis of exosomal mRNAs in human cerebrospinal fluid. BMC Med Genet 12:7
Caradec J, Kharmate G, Hosseini-Beheshti E et al (2014) Reproducibility and efficiency of serum-derived exosome extraction methods. Clin Biochem 47:1286–1292
Lässer C, Alikhani VS, Ekström K et al (2011) Human saliva, plasma and breast milk exosomes contain RNA: uptake by macrophages. J Transl Med 9(9)
Zhou H, Yuen PST, Pisitkun T et al (2006) Collection, storage, preservation, and normalization of human urinary exosomes for biomarker discovery. Kidney Int 69(8):1471–1476
Lo CA, Delevoye C, Gilles-Marsens F et al (2015) Exosomes released by keratinocytes modulate melanocyte pigmentation. Nat Commun 6:7506
Kotzerke K, Mempel M, Aung T et al (2013) Immunostimulatory activity of murine keratinocyte-derived exosomes. Exp Dermatol 22:650–655
Sjoqvist S, Kasai Y, Shimura D et al (2019) Oral keratinocyte-derived exosomes regulate proliferation of fibroblasts and epithelial cells. Biochem Biophys Res Commun 514:706–712
Sjöqvist S, Ishikawa T, Shimura D et al (2019) Exosomes derived from clinical-grade oral mucosal epithelial cell sheets promote wound healing. J Extracell Vesicles 8:1565264
Lobb RJ, Becker M, Wen SW et al (2015) Optimized exosome isolation protocol for cell culture supernatant and human plasma. J Extracell Vesicles 1:1–11
Nordin JZ, Lee Y, Vader P et al (2015) Ultrafiltration with size-exclusion liquid chromatography for high yield isolation of extracellular vesicles preserving intact biophysical and functional properties. Nanomedicine 11:1–5
Stranska R, Gysbrechts L, Wouters J et al (2018) Comparison of membrane affinity-based method with size-exclusion chromatography for isolation of exosome-like vesicles from human plasma. J Transl Med 16:1
Théry C, Witwer KW, Aikawa E et al (2018) 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(1):1535750
Corso G, Mäger I, Lee Y et al (2017) Reproducible and scalable purification of extracellular vesicles using combined bind-elute and size exclusion chromatography. Sci Rep 7:11561
Acknowledgments
This work was supported by the Swedish Society of Medicine. SELA is supported by the Swedish Research Council (VR-Med) and the Swedish foundation of Medical Research (SSF-IRC).
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media New York
About this protocol
Cite this protocol
Sjöqvist, S., Imafuku, A., Gupta, D., EL Andaloussi, S. (2019). Isolation and Characterization of Extracellular Vesicles from Keratinocyte Cultures. In: Turksen, K. (eds) Epidermal Cells. Methods in Molecular Biology, vol 2109. Humana, New York, NY. https://doi.org/10.1007/7651_2019_264
Download citation
DOI: https://doi.org/10.1007/7651_2019_264
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0250-8
Online ISBN: 978-1-0716-0251-5
eBook Packages: Springer Protocols