A Single Extracellular Vesicle (EV) Flow Cytometry Approach to Reveal EV Heterogeneity

Abstract

Extracellular vesicles (EVs) actively participate in intercellular communication and pathological processes. Studying the molecular signatures of EVs is key to reveal their biological functions and clinical values,w hich, however, is greatly hindered by their sub-100 nm dimensions,t he low quantities of biomolecules each EV carries,a nd the large population heterogeneity.N ow,s ingle-EV flow cytometry analysis is introduced to realizes ingle EV counting and phenotyping in aconventional flowcytometer for the first time, enabled by target-initiated engineering (TIE) of DNAn anostructures on each EV.B yi lluminating multiple markers on single EVs,s tatistically significant differences are revealed among the molecular signatures of EVs originating from several breast cancer cell lines,and the cancer cell-derived EVs among the heterogeneous EV populations are successfully recognized. Thus,o ur approachh olds great potential for various biological and biomedical applications.Supportinginformation and the ORCID identification number(s) for the author(s) of this article can be found under: https://doi.

An immunogold single extracellular vesicular RNA and protein (AuSERP) biochip to predict responses to immunotherapy in non‐small cell lung cancer patients

“…Therefore, it is imperative to develop technologies that provide an accurate and efficient analysis of the molecular content at a single‐EV level. Several single‐EV analytical technologies have been proposed for EV molecular analysis, including analysis by fluorescence imaging of immobilized single EVs (Fraser et al., 2019; Lee et al., 2018; Liu et al., 2019), flow cytometry of single EVs via target‐initiated engineering (Shen et al., 2018), droplet‐based single‐exosome‐counting ELISA (droplet digital ExoELISA) (Liu et al., 2018), digital detection integrated with surface‐anchored nucleic acid amplification (Tian et al., 2018), proximity‐dependent barcoding assay (Wu et al., 2019), and immuno‐droplet digital PCR (iddPCR) (Ko et al., 2020). These methods successfully improved the limit of detection (LOD) and demonstrated heterogeneous protein profiles of single EVs unachievable by bulk‐analysis methods.…”

Mixed-Dimensional Heterostructure Material-Based SERS for Trace Level Identification of Breast Cancer-Derived Exosomes

“… 40 − 49 Because exosomes can be obtained easily from biological fluids in clinics, they have been considered as minimally invasive cancer biomarkers. 38 − 43 …”

Extracellular vesicles in type 2 diabetes mellitus: key roles in pathogenesis, complications, and therapy

“…Furthermore, single‐EV flow cytometry could realize single EV counting and phenotyping by target‐initiated engineering of DNA nanostructures on each EV. By identifying markers on single EVs, single‐EVs flow cytometry could efficiently recognize cancer cell‐derived EVs among heterogeneous EVs populations [40]. Therefore, flow cytometry may be an effective tool for the analysis of EVs.…”