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. 2023:2718:253-269.
doi: 10.1007/978-1-0716-3457-8_14.

Density-Based Fractionation of Cell-Conditioned Medium to Prepare Proteomics Grade Extracellular Vesicles

Quentin Roux  1   2 , Sarah Deville  1   2 , An Hendrix  3   4
Affiliations

Density-Based Fractionation of Cell-Conditioned Medium to Prepare Proteomics Grade Extracellular Vesicles

Quentin Roux et al. Methods Mol Biol. 2023.

Abstract

The identification of the molecular composition of extracellular vesicles (EV) by omics approaches, including proteomics, requires the separation of EV from non-EV confounding factors present in the source biofluid. In this protocol, we present the sequential implementation of density gradient ultracentrifugation and size-exclusion chromatography to prepare EV from cell-conditioned medium with high specificity and repeatability. This approach enables the recovery of intact purified EV suited for downstream functional assays and biomarker discovery by omics approaches.

Keywords: Cell culture; Density gradient; Extracellular vesicles; Proteomics; Size-exclusion chromatography.

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