Rapid isolation of small extracellular vesicles by oligonucleotide functionalized magnetic beads and detection their surface proteins using DARPin probe
Alexey M. Yashchenok1, Vasiliy S. Chernyshev1, Elena V. Konovalova2, Roman Kholodenko2, Ekaterina Tsydenzhapova3, Viktoria O. Shipunova3, Alexey A. Schulga2, Sergey M. Deyev2, Dmitry A. Gorin1
1 Center for Photonic Science and Engineering, Skolkovo Institute of Science and Technology, Moscow, Russia
2 Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
3 Moscow Institute of Physics and Technology, Moscow Region, Russia
Abstract
Small extracellular vesicles (sEVs) are a type of membrane nanocarriers that carry molecular cargo of parental cells. In addition to their exceptional role in cellular functions and communication, sEVs are a promising biomarker for diagnosing diseases in the very early stages, such as cancer and neurodegenerative disorders. In this regard, the development of a sensitive and reliable method for the isolation and detection of pathologically relevant sEVs remains a challenge. In this study, we present a method for rapid characterization of sEVs that is based on the isolation of sEVs by anti-CD63 aptamer conjugated magnetic beads followed by detecting the presence of the EpCAM and HER2 proteins on the surface of sEVs by anti-EpCAM and anti-HER2 designed ankyrin repeat proteins (DARPins). By combining magnetic capture and fluorescence identification, the method enables highly sensitive (~ 10^4 sEVs per mL), rapid (1 hour) and specific sEVs detection. This method provides a basis for isolating and quantifying various sEV subpopulations that contain disease specific surface-markers.
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Speaker
Alexey Yashchenok
Center for Photonic Science and Engineering, Skolkovo Institute of Science and Technology, Moscow, Russia
Russia
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