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Multimodal Analytical Platform on a Multiplexed Surface Plasmon Resonance Imaging Chip for the Analysis of Extracellular Vesicle Subsets
Published on: March 17, 2023
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A Combined Western and Bead-Based Multiplex Platform to Characterize Extracellular Vesicles
Josette C van Maanen1, Frances C Bach1, Theresa S Braun2
1Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, the Netherlands.
Tissue Engineering. Part C, Methods
|July 20, 2023
Summary
This study introduces DigiWest, a multiplex platform for extracellular vesicle (EV) characterization. It efficiently analyzes multiple EV markers with low protein input, aiding regenerative medicine research.
Area of Science:
- Regenerative Medicine
- Cell Biology
- Biochemistry
Background:
- Extracellular vesicles (EVs) are crucial for intercellular communication and show promise in regenerative medicine due to their low immunogenicity.
- Current methods for EV characterization, like Western blotting, require substantial protein input for analyzing individual markers.
- International Society for Extracellular Vesicles (ISEV) guidelines recommend analyzing multiple EV markers and co-isolated proteins to ensure purity and composition.
Purpose of the Study:
- To evaluate the DigiWest platform for simultaneous detection of multiple EV markers.
- To assess the efficiency of DigiWest in terms of protein input compared to traditional Western blotting.
- To establish a cross-species antibody panel for comprehensive EV characterization.
Main Methods:
- DigiWest, a combined Western and bead-based multiplex platform, was used for EV analysis.
- EVs from human, pig, and dog sources (mesenchymal stromal cells, notochordal cells, milk) were analyzed.
- A panel of nine antibodies was developed for detecting general EV markers and co-isolates.
Main Results:
- DigiWest enabled simultaneous detection of multiple EV markers with significantly lower protein input (1 μg) compared to Western blotting (∼10 μg per marker).
- The study successfully established a cross-species antibody panel for EV marker analysis.
- The platform demonstrated effectiveness across various EV sources and species.
Conclusions:
- The DigiWest platform offers a highly efficient method for comprehensive EV characterization.
- This technique facilitates adherence to ISEV guidelines for EV purity and composition analysis.
- DigiWest has significant potential for advancing research and applications in regenerative medicine.
