CRISPR/Cas9 Genome Editing vs. Over-Expression for Fluorescent Extracellular Vesicle-Labeling: A Quantitative Analysis.

ABSTRACT: Over-expression of fluorescently-labeled markers for extracellular vesicles is frequently used to visualize vesicle up-take and transport. EVs that are labeled by over-expression show considerable heterogeneity regarding the number of fluorophores on single particles, which could potentially bias tracking and up-take studies in favor of more strongly-labeled particles. To avoid the potential artefacts that are caused by over-expression, we developed a genome editing approach for the fluorescent labeling of the extracellular vesicle marker CD63 with green fluorescent protein using the CRISPR/Cas9 technology. Using single-molecule sensitive fluorescence microscopy, we quantitatively compared the degree of labeling of secreted small extracellular vesicles from conventional over-expression and

SUBMITTER: Strohmeier K 

PROVIDER: S-EPMC8745383 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature