Aptasensor-enabled quantitative analysis of nano-sized extracellular vesicles by flow cytometry

Abstract

Extracellular vesicles (EVs) represent an important mode of intercellular communication in both disease and developmental biology, exposing their potential in diagnostics and therapeutics. Recently, aptamer-based sensors, i.e. aptasensors, have been gradually applied in EV analysis due to their high selectivity and sensitivity. A fluorescent aptasensor enables easy readout by flow cytometry (FCM) and has more accuracy and convenience than conventional immunoassays for EV analysis. Here, we develop a fluorescent aptasensor-based method for quantitative analysis of nano-sized membrane vesicles by using high-resolution FCM. EVs as small as 100 nm are detected and quantified using a dual-staining procedure with the fluorescent aptasensor targeting CD63 and a cytoplasmic dye. Nano-sized EVs derived from bone marrow mesenchymal stem cells, human neural stem cells and human cornea epithelial cells are analyzed, and the result shows that their amount varies from 6.79 × 10⁶ mL⁻¹ to 2.08 × 10⁸ mL⁻¹ in culture media. The technique is also used to evaluate the bioactivity of EVs and, in the future, it may develop into a versatile tool to analyze and quantify EVs from a variety of biological objects with conventional cytometric instruments.