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Published on: January 7, 2019
High Purity Isolation and Sensitive Quantification of Extracellular Vesicles Using Affinity to TIM4
Takeshi Yoshida1, Takamasa Ishidome1, Rikinari Hanayama1
1Department of Immunology, Kanazawa University Graduate School of Medical Sciences, Kanazawa, Japan.
Abstract:
Almost all types of cells secrete extracellular vesicles (EVs), including exosomes and microvesicles. EVs carry various proteins, lipids, mRNAs, and microRNAs, and may participate in many aspects of physiological and pathophysiological processes. Various studies are currently being conducted to develop therapeutic and diagnostic methods targeting or utilizing EVs. Therefore, developing ideal methods for isolating and quantifying EVs is an active area of research. EVs express phosphatidylserine on their outer lipid bilayer. This unit describes an affinity-based method for isolating EVs using TIM4, which binds phosphatidylserine in a specific and calcium-dependent manner. EVs captured by TIM4 can be easily released by addition of a chelating agent, or can be retained for quantification by ELISA or flow cytometry. These methods enable the isolation of highly purified EVs and the sensitive quantification of EVs, which will accelerate EV research beyond what has been achievable with conventional methods. © 2017 by John Wiley & Sons, Inc.
Insights
Researchers developed a new method using TIM4 to isolate and quantify extracellular vesicles (EVs). This affinity-based technique improves purification and detection sensitivity, advancing EV research for therapeutic and diagnostic applications.
Area of Science:
- Biotechnology
- Cell Biology
- Biochemistry
Background:
- Extracellular vesicles (EVs), including exosomes and microvesicles, are secreted by most cells and play roles in physiological and pathophysiological processes.
- Current research focuses on developing therapeutic and diagnostic strategies involving EVs, necessitating improved isolation and quantification methods.
- EVs are characterized by the presence of phosphatidylserine on their outer membrane.
Purpose of the Study:
- To describe an affinity-based method for isolating and quantifying extracellular vesicles (EVs).
- To utilize the phosphatidylserine-binding protein TIM4 for specific and efficient EV capture.
- To enable sensitive quantification of EVs for accelerated research.
Main Methods:
- An affinity-based isolation technique using TIM4, a protein that binds phosphatidylserine on EVs.
- EVs captured by TIM4 can be released using a chelating agent.
- Quantification of captured EVs via ELISA or flow cytometry.
Main Results:
- The TIM4-based method allows for the isolation of highly purified EVs.
- The method enables sensitive quantification of EVs.
- This approach facilitates EV research beyond conventional techniques.
Conclusions:
- The described TIM4 affinity method provides a robust approach for EV isolation and quantification.
- This technique enhances the purity and sensitivity of EV analysis.
- The findings are expected to accelerate the development of EV-based diagnostics and therapeutics.
