Profiling extracellular vesicle surface proteins with 10 µL peripheral plasma within 4 h

Area of Science:

• Biochemistry
• Molecular Biology
• Nanotechnology

Background:

• Extracellular vesicle (EV) surface proteins are crucial biomarkers for early cancer detection.
• Detecting these EV proteins is challenging due to low abundance and interference in clinical samples.

Purpose of the Study:

• To develop a sensitive and rapid method for direct detection of EV surface proteins.
• To establish a diagnostic tool for analyzing cancer biomarkers on EVs.

Main Methods:

• A MEmbrane-Specific Separation and two-step Cascade AmpLificatioN (MESS2CAN) strategy was developed.
• Utilized novel lipid probes and streptavidin-coated magnetic beads for EV recovery (49.6% in 1 hour).
• Employed a dual amplification strategy (primer exchange reaction cascaded by Cas12a) for sensitive protein detection (10 EV particles/µL).

Main Results:

• MESS2CAN achieved high EV recovery and sensitive detection of target proteins.
• Demonstrated successful analysis of HER2, EpCAM, and EGFR on EVs from cell lines and patient plasma.
• Achieved high specificity and sensitivity for EGFR (AUC=0.98) and HER2 (AUC=1) in discriminating breast cancer subtypes and healthy donors.

Conclusions:

• MESS2CAN is a pioneering method for highly sensitive in vitro EV diagnostics.
• The method is applicable to clinical samples with trace amounts of EVs.
• MESS2CAN offers a promising approach for early cancer diagnosis through EV surface protein analysis.