Simple and rapid extracellular vesicles quantification via membrane biotinylation strategy coupled with fluorescent nanospheres-based lateral flow assay.

Extracellular vesicles (EVs) are cell-excreted membrane particles existing in a variety of biological fluids. As potential noninvasive biomarkers, EVs have received wide attention in recent years. However, usual EVs assays are complex, time-consuming and costly, thus limiting their clinical utility. Simple and rapid EVs quantification within biological fluids remains challenging. Here, we developed a simple, rapid strategy for EVs quantification, which combined with lateral flow assay and membrane biotinylation strategy. By utilizing biotin-functionalized phosphatidylethanolamine (DSPE-PEG-Biotin), the membrane of EVs could be successfully modified with biotin under strong hydrophobic interactions. Subsequently, based on the high affinity between streptavidin and biotin, quantification assay was achieved by lateral flow assay with fluorescent nanospheres (FNs) as a reporter. Biotinylation of biogenic EVs could be reached to 85%. This proposed method enables sensitive detection of 2.0 × 103 particles/μL. The whole procedure time was within 1 h. Furthermore, this approach was used to detect EVs in biological samples, demonstrating potential clinical applications.