Colao, Ivano Luigi;
Corteling, Randolph L;
Bracewell, Daniel G;
Wall, Ivan B;
(2025)
Neural stem cell–derived extracellular vesicles purified by monolith chromatography retain stimulatory effect in in vitro scratch assay.
Cytotherapy
, 27
(3)
pp. 365-377.
10.1016/j.jcyt.2024.11.007.
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Abstract
Background aims: Extracellular vesicles (EVs) have gained traction as potential cell-free therapeutic candidates. Development of purification methods that are scalable and robust is a major focus of EV research. Yet there is still little in the literature that evaluates purification methods against potency of the EV product. In the present study, we examined two monolith chromatography methods with a focus on assessing the ability of purified EVs to retain stimulatory effects on fibroblasts to connect scalable purification methods with product outputs. // Methods: We characterized EVs recovered from CTX0E03 (CTX) neural stem cell–conditioned medium in terms of biomarker distribution, functional capacity and purity. We evaluated the ability of EVs to promote wound closure in an in vitro scratch assay prior to and following two monolith chromatography steps (anion exchange and hydrophobic interaction) to determine whether these options may better serve EV bioprocessing. // Results: EVs from CTX cells were successful in initiating wound repair in a fibroblast scratch assay over 72 h with a single 20-μg dose. EV preparations presented the markers CD9, CD81 and CD63 but also contained culture albumin and DNA as process impurities. EVs recovered by tangential flow filtration could be successfully purified further by both monolith chromatography steps. Post-monolith EV stimulation was conserved. // Conclusions: The results indicate that monolith chromatography is a viable purification method for EVs derived from cell culture that does not detract from the product's ability to stimulate fibroblasts, suggesting that product functionality is conserved. Further work is needed in developing suitable downstream processes and analytics to achieve clinically relevant purities for injectable biologics.
| Type: | Article |
|---|---|
| Title: | Neural stem cell–derived extracellular vesicles purified by monolith chromatography retain stimulatory effect in in vitro scratch assay |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1016/j.jcyt.2024.11.007 |
| Publisher version: | http://dx.doi.org/10.1016/j.jcyt.2024.11.007 |
| Language: | English |
| Additional information: | Copyright © 2025 International Society for Cell & Gene Therapy. Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
| Keywords: | Extracellular vesicles; hydrophobic interaction; neural stem cells; quaternary amine; scratch assay; TFF |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > UCL BEAMS UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10200835 |
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