Affiliation:
1. Wyss Institute for Biologically Inspired Engineering
2. Tufts University School of Medicine
3. Department of Pathology, Brigham and Women’s Hospital
4. Harvard Medical School
Abstract
Extracellular vesicles (EVs) are released by all cells into biofluids and hold great promise as reservoirs of disease biomarkers. One of the main challenges in studying EVs is a lack of methods to quantify EVs that are sensitive enough and can differentiate EVs from similarly sized lipoproteins and protein aggregates. We demonstrate the use of ultrasensitive, single-molecule array (Simoa) assays for the quantification of EVs using three widely expressed transmembrane proteins: the tetraspanins CD9, CD63, and CD81. Using Simoa to measure these three EV markers, as well as albumin to measure protein contamination, we were able to compare the relative efficiency and purity of several commonly used EV isolation methods in plasma and cerebrospinal fluid (CSF): ultracentrifugation, precipitation, and size exclusion chromatography (SEC). We further used these assays, all on one platform, to improve SEC isolation from plasma and CSF. Our results highlight the utility of quantifying EV proteins using Simoa and provide a rapid framework for comparing and improving EV isolation methods from biofluids.
Funder
Chan Zuckerberg Initiative
Open Philanthropy Project
Publisher
eLife Sciences Publications, Ltd
Subject
General Immunology and Microbiology,General Biochemistry, Genetics and Molecular Biology,General Medicine,General Neuroscience
Cited by
45 articles.
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