Sensitive and Specific Detection of Platelet-Derived and Tissue Factor–Positive Extracellular Vesicles in Plasma Using Solid-Phase Proximity Ligation Assay

• Main Authors: Åsa Thulin, Junhong Yan, Mikael Åberg, Christina Christersson, Masood Kamali-Moghaddam, Agneta Siegbahn
• Format: Article
• Language: English
• Published: Georg Thieme Verlag KG 2018-07-01
• Series: TH Open
• Subjects: microvesicles; myocardial infarction; cardiovascular diseases; vascular homeostasis
• Online Access: http://www.thieme-connect.de/DOI/DOI?10.1055/s-0038-1667204
• ISSN: 2512-9465; 2512-9465

Extracellular vesicles (EVs) derived from blood cells are promising biomarkers for various diseases. However, they are difficult to measure accurately in plasma due to their small size. Here, we demonstrate that platelet-derived EVs in plasma can be measured using solid-phase proximity ligation assay with high sensitivity and specificity using very small sample volume of biological materials. The results correlate well with high-sensitivity flow cytometry with the difference that the smallest EVs are detected. Briefly, the EVs are first captured on a solid phase, using lactadherin binding, and detection requires recognition with two antibodies followed by qPCR. The assay, using cholera toxin subunit-B or lactadherin as capture agents, also allowed detection of the more rare population of tissue factor (TF)-positive EVs at a concentration similar to sensitive TF activity assays. Thus, this assay can detect different types of EVs with high specificity and sensitivity, and has the potential to be an attractive alternative to flow cytometric analysis of preclinical and clinical samples. Improved techniques for measuring EVs in plasma will hopefully contribute to the understanding of their role in several diseases.