Abstract
Extracellular vesicles (EVs) are secreted by eukaryotic cells and serve as carriers for a variety of cell signaling factors, including RNAs, proteins, and lipids. We described a unique population of EVs, the membrane of which is highly enriched with the sphingolipid ceramide. We suggested that ceramide in the EV membrane is organized in ceramide-rich platforms (CRPs), a type of lipid raft that mediates interaction of ceramide with ceramide-associated proteins (CAPs). Here, we describe methods using anti-ceramide antibody to isolate ceramide-enriched EVs and detect exosomes after uptake into recipient cells. In addition, we discuss methods for EV analysis using nanoparticle tracking and mass spectrometry. The methods can be extended to the isolation of other types of EVs and “mobile rafts” transported by EVs from donor to recipient cells using antibodies against lipids specific for these EVs.
Original language | English |
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Title of host publication | Methods in Molecular Biology |
Pages | 87-98 |
Number of pages | 12 |
DOIs | |
State | Published - 2021 |
Publication series
Name | Methods in Molecular Biology |
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Volume | 2187 |
ISSN (Print) | 1064-3745 |
ISSN (Electronic) | 1940-6029 |
Bibliographical note
Publisher Copyright:© 2021, Springer Science+Business Media, LLC, part of Springer Nature.
Funding
This work was in part supported by NIH grants R01AG034389 and R01AG064234, and Department of Veteran Affairs grant I01 BX003643. The authors are grateful for support by the Department of Physiology (Chair Dr. Alan Daugherty) at the University of Kentucky, Lexington, KY.
Funders | Funder number |
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Department of Veteran Affairs | I01 BX003643 |
National Institutes of Health (NIH) | R01AG034389 |
National Institutes of Health (NIH) | |
National Institute on Aging | R01AG064234 |
National Institute on Aging |
Keywords
- Affinity chromatography
- Alzheimer’s disease
- Ceramide-rich platform
- Extracellular vesicles
- Sphingolipid
ASJC Scopus subject areas
- Molecular Biology
- Genetics