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

Jie He; Hengyu Li; John Mai; Yuqing Ke; Chunhui Zhai; Jiao Jiao Li; Lai Jiang; Guangxia Shen; Xianting Ding

doi:10.1002/jev2.12364

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

J Extracell Vesicles. 2023 Sep;12(9):e12364. doi: 10.1002/jev2.12364.

Authors

Jie He^{1

2}, Hengyu Li³, John Mai⁴, Yuqing Ke^{1

2}, Chunhui Zhai^{1

2}, Jiao Jiao Li⁵, Lai Jiang^{1

2}, Guangxia Shen^{1

2}, Xianting Ding^{1

2}

Affiliations

¹ Department of Anesthesiology and Surgical Intensive Care Unit School of Medicine and School of Biomedical Engineering, Xinhua Hospital, Shanghai Jiao Tong University, Shanghai, China.
² State Key Laboratory of Oncogenes and Related Genes, Institute for Personalized Medicine, Shanghai Jiao Tong University, Shanghai, China.
³ Department of Breast and Thyroid Surgery, Changhai Hospital, Naval Military Medical University, Shanghai, China.
⁴ Alfred E. Mann Institute for Biomedical Engineering, University of Southern California, Los Angeles, California, USA.
⁵ School of Biomedical Engineering Faculty of Engineering and IT, University of Technology Sydney, Sydney, NSW, Australia.

Abstract

Extracellular vesicle (EV) surface proteins, expressed by primary tumours, are important biomarkers for early cancer diagnosis. However, the detection of these EV proteins is complicated by their low abundance and interference from non-EV components in clinical samples. Herein, we present a MEmbrane-Specific Separation and two-step Cascade AmpLificatioN (MESS2CAN) strategy for direct detection of EV surface proteins within 4 h. MESS2CAN utilises novel lipid probes (long chains linked by PEG2K with biotin at one end, and DSPE at the other end) and streptavidin-coated magnetic beads, permitting a 49.6% EV recovery rate within 1 h. A dual amplification strategy with a primer exchange reaction (PER) cascaded by the Cas12a system then allows sensitive detection of the target protein at 10 EV particles per microliter. Using 4 cell lines and 90 clinical test samples, we demonstrate MESS2CAN for analysing HER2, EpCAM and EGFR expression on EVs derived from cells and patient plasma. MESS2CAN reports the desired specificity and sensitivity of EGFR (AUC = 0.98) and of HER2 (AUC = 1) for discriminating between HER2-positive breast cancer, triple-negative breast cancer and healthy donors. MESS2CAN is a pioneering method for highly sensitive in vitro EV diagnostics, applicable to clinical samples with trace amounts of EVs.

Keywords: breast cancer; extracellular vesicles; in vitro diagnostics; protein analysis.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Biotin
Breast Neoplasms* / diagnosis
ErbB Receptors
Extracellular Vesicles*
Female
Humans
Membrane Proteins

Substances

Membrane Proteins
Biotin
ErbB Receptors