Extracellular-Vesicle Catch-and-Release Isolation System Using a Net-Charge Invertible Curvature-Sensing Peptide

Area of Science:

• Biotechnology
• Cell Biology
• Biochemistry

Background:

• Extracellular vesicles (EVs) are crucial for intercellular communication, carrying diverse biomolecules.
• EVs are promising diagnostic and therapeutic targets in medicine.
• Current EV isolation methods, like ultracentrifugation, are equipment-intensive and challenging.

Purpose of the Study:

• To develop a novel, simple, and convenient method for isolating extracellular vesicles (EVs).
• To introduce a pH-dependent peptide for reversible EV capture and release.
• To enable efficient EV isolation for downstream applications, including single-particle analysis.

Main Methods:

• Development of an EV catch-and-release isolation system (EV-CaRiS).
• Design of a net-charge invertible curvature-sensing peptide (NIC) for EV binding.
• Application of a batch isolation method using NIC-functionalized resin.
• Utilizing total internal reflection fluorescence microscopy (TIRFM) for single-particle imaging.

Main Results:

• Reproducible isolation of EVs from three human cell lines was achieved using EV-CaRiS.
• NIC successfully captured and released EVs in a pH-dependent manner.
• Single-particle imaging confirmed the presence of exosome markers (CD63, CD81) in isolated EVs.
• The method demonstrated simplicity and convenience compared to traditional techniques.

Conclusions:

• EV-CaRiS provides a simple and convenient methodology for extracellular vesicle isolation.
• The NIC peptide is a promising tool for EV isolation and single-particle analysis.
• This novel system facilitates research in EV biology and clinical applications.