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Volume 26
Issue 17
10.3390/ijms26178477
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This is an early access version, the complete PDF, HTML, and XML versions will be available soon.
Article

Comprehensive Characterisation of Extracellular Vesicle Preparations Using Multiparametric Size-Exclusion Chromatography

by
Darja Božič
*,
Katja Vrabec
,
Ana Železnik
,
Andrej Raspor
,
Valentina Novak
,
Ivana Petrović Koshmak
,
Maja Leskovec
and
Aleš Štrancar
Sartorius Bia Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2025, 26(17), 8477; https://doi.org/10.3390/ijms26178477 (registering DOI)
Submission received: 14 July 2025 / Revised: 8 August 2025 / Accepted: 27 August 2025 / Published: 31 August 2025
(This article belongs to the Special Issue Recent Progress in Extracellular Vesicles)
Versions Notes

Abstract

Extracellular vesicles (EVs) hold great promise in the fields of diagnostics and therapeutics. However, the heterogeneity of these membrane-enclosed messengers and the complexity of the biological samples in which they occur pose significant research challenges. The aim of this study was to improve the reliability of size-exclusion chromatography (SEC) and immunolabelling as common approaches in the EV field, and to provide a comprehensive characterisation tool for diverse EV preparations. Profiling of SEC-separated sample components was conducted through light absorbance, fluorescence, and light scattering, providing insights into particle content, size, protein content, and specific markers. Key considerations for assay robustness, including SEC mobile phase composition and immunostaining parameters, were addressed. Respecting the importance of controlled immunolabelling and preanalytical factors, the method efficiently reveals changes in the sample profile with respect to particles and small impurities. The detailed analytical capabilities and method adaptability offer a practical way to enhance the efficiency of EV research and applications.
Keywords: extracellular vesicles; analytical chromatography; fluorescence; MALS; PAT; immunolabelling; quality control extracellular vesicles; analytical chromatography; fluorescence; MALS; PAT; immunolabelling; quality control

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MDPI and ACS Style

Božič, D.; Vrabec, K.; Železnik, A.; Raspor, A.; Novak, V.; Petrović Koshmak, I.; Leskovec, M.; Štrancar, A. Comprehensive Characterisation of Extracellular Vesicle Preparations Using Multiparametric Size-Exclusion Chromatography. Int. J. Mol. Sci. 2025, 26, 8477. https://doi.org/10.3390/ijms26178477

AMA Style

Božič D, Vrabec K, Železnik A, Raspor A, Novak V, Petrović Koshmak I, Leskovec M, Štrancar A. Comprehensive Characterisation of Extracellular Vesicle Preparations Using Multiparametric Size-Exclusion Chromatography. International Journal of Molecular Sciences. 2025; 26(17):8477. https://doi.org/10.3390/ijms26178477

Chicago/Turabian Style

Božič, Darja, Katja Vrabec, Ana Železnik, Andrej Raspor, Valentina Novak, Ivana Petrović Koshmak, Maja Leskovec, and Aleš Štrancar. 2025. "Comprehensive Characterisation of Extracellular Vesicle Preparations Using Multiparametric Size-Exclusion Chromatography" International Journal of Molecular Sciences 26, no. 17: 8477. https://doi.org/10.3390/ijms26178477

APA Style

Božič, D., Vrabec, K., Železnik, A., Raspor, A., Novak, V., Petrović Koshmak, I., Leskovec, M., & Štrancar, A. (2025). Comprehensive Characterisation of Extracellular Vesicle Preparations Using Multiparametric Size-Exclusion Chromatography. International Journal of Molecular Sciences, 26(17), 8477. https://doi.org/10.3390/ijms26178477

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