Abstract

High-Resolution Analysis of Single Extracellular Vesicles and Particles with Digital Flow Cytometry and Super-Resolution Imaging

Daniel Chiu, Professor, University of Washington

We have developed a multi-parametric high-throughput flow-based method for the analysis of individual extracelluar vesicles and particles (EVPs), and a super-resolution method for sizing individual EVPs in a high-throughput fashion. EVPs are highly heterogeneous and comprise a diverse set of surface protein markers as well as intra-vesicular cargoes. Yet, current approaches to the study of EVPs lack the necessary sensitivity and precision to fully characterize and understand the make-up and the distribution of various EV subpopulations that may be present. Digital flow cytometry (dFC) provides single-fluorophore sensitivity and enables multiparameter characterization of EVPs, including sin­gle-EVP phenotyping, the absolute quantitation of EVP concentrations, and biomarker copy numbers. dFC has a broad range of applications, from analysis of single EVPs such as exosomes or RNA-binding proteins to characterization of therapeutic lipid nano­particles, viruses, and proteins. dFC also provides absolute quantita­tion of non-EVP samples such as dyes, beads, and Ab-dye conjugates.