2023
DOI: 10.1021/acs.analchem.3c00144
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Purification Analysis, Intracellular Tracking, and Colocalization of Extracellular Vesicles Using Atomic Force and 3D Single-Molecule Localization Microscopy

Abstract: Extracellular vesicles (EVs) play a key role in cell− cell communication and thus have great potential to be utilized as therapeutic agents and diagnostic tools. In this study, we implemented single-molecule microscopy techniques as a toolbox for a comprehensive characterization as well as measurement of the cellular uptake of HEK293T cell-derived EVs (eGFP-labeled) in HeLa cells. A combination of fluorescence and atomic force microscopy revealed a fraction of 68% fluorescently labeled EVs with an average size… Show more

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“…This holds promise for the development of more biocompatible and scalable intracellular drug delivery nanocarriers with a reduced immunogenicity. These results align well with previous studies utilizing Nanoflow cytometry , and single molecule localization microscopy, which have also observed a similar range of cargo loading in engineered EV subpopulations, with a similar percentage of cargo-loaded extracellular vesicles at the single-molecule and single-vesicle level. Furthermore, the additional cleaning procedures in this study provided stable results for Ab-loading and insights into the relationship between nanovesicle physical properties and cargo nature, offering opportunities to enhance the purity of production yields and customize these nanovesicles for specific therapeutic agent for Ab delivery with substantially lower side effects.…”
Section: Discussionmentioning
confidence: 99%
“…This holds promise for the development of more biocompatible and scalable intracellular drug delivery nanocarriers with a reduced immunogenicity. These results align well with previous studies utilizing Nanoflow cytometry , and single molecule localization microscopy, which have also observed a similar range of cargo loading in engineered EV subpopulations, with a similar percentage of cargo-loaded extracellular vesicles at the single-molecule and single-vesicle level. Furthermore, the additional cleaning procedures in this study provided stable results for Ab-loading and insights into the relationship between nanovesicle physical properties and cargo nature, offering opportunities to enhance the purity of production yields and customize these nanovesicles for specific therapeutic agent for Ab delivery with substantially lower side effects.…”
Section: Discussionmentioning
confidence: 99%