Size Exclusion Chromatography: Optimizing Extracellular Vesicle Isolation

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Size Exclusion Chromatography: The Key for Optimizing Extracellular Vesicle Isolation

When it comes to EVs purification, Size Exclusion Chromatography (SEC) presents multiple advantages compared to other methods.

Extracellular vesicles (EVs) are quickly gaining more interest within Healthcare Industry since they have enormous medical and biotechnological potential as diagnostic tools and treatment vehicles against diseases like cancer.

But, what is the nature of EVs? EVs are lipid-bound sacs secreted by cells into the extracellular space whose biological action is to transfer information to other cells, thereby influencing the recipient cell function.

However, one of the main challenges is this field is the EVs isolation and purification. EVs are extracted from complex biofluid samples which contain soluble proteins and lipoproteins (LLPs) as well.

Therefore, finding an adequate isolation method that results in high performance, purity, ease of handling, cost-effectiveness and shorter timing is essential.

Figure 1. Graphical abstract of the extracellular vesicle isolation process


When it comes to EVs purification, Size Exclusion Chromatography (SEC) using agarose resins from Agarose Bead Technologies (ABT) presents multiple advantages compared to other methods.

SEC eliminates soluble proteins, provides higher yields, does not require specific markers for EV collection, and collects all EVs without subpopulation enrichment.

SEC is cost-effective, easily adaptable for sample processing, and does not require expensive equipment. It separates particles based on their size, allowing EVs to pass through the column to elute earlier while retaining smaller particles.

At ABT, the agarose resin characteristics can be customized. For instance, for EVs coming from Human melanoma cell line SK-MEL-147 and blood samples from healthy donors, parameters like crosslinking level, bead, and pore size are determinants to obtain an optimal result in terms of stability, resolution, and exclusion limit.

In this case, reduced bead size and the selection of a 4% of agarose resin greatly improved EV purity and yield. Demonstrated not only by immunodetection and absorbance measurements of EVs and LPPs levels but by nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM).

Consequently, it has been shown that the use of 4% Rapid Run Fine agarose bead resin* from ABT is better than the gold standard method both on a lab scale and on a large scale for EVs isolation and purification.


1. Yáñez-Mó, M. et al. (2015). Biological properties of extracellular vesicles and their physiological functions. Journal of Extracellular Vesicles, 4, 27066.

2. Benayas, B., Morales, J., Egea, C., Armisén, P., & Yáñez-Mó, M. (2023). Optimization of extracellular vesicle isolation and their separation from lipoproteins by size exclusion chromatography. Journal of Extracellular Biology, 2, e100.

* Customized product.

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