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Article

Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine

by
Julia Neumair
1,
Claudia D’Ercole
2,
Matteo De March
2,
Martin Elsner
1,
Michael Seidel
1 and
Ario de Marco
2,*
1
Chair of Analytical Chemistry and Water Chemistry, Technical University of Munich, Lichtenbergstr. 4, 85748 Garching, Germany
2
Laboratory of Environmental and Life Sciences, University of Nova Gorica, Vipavska Cesta 13, P.O. Box 301, SI-5000 Nova Gorica, Slovenia
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2023, 24(7), 6131; https://doi.org/10.3390/ijms24076131
Submission received: 21 February 2023 / Revised: 17 March 2023 / Accepted: 21 March 2023 / Published: 24 March 2023
(This article belongs to the Special Issue Advances in Single Domain-Based Antibodies)
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Abstract

Extracellular vesicles (EVs) have enormous potential for the implementation of liquid biopsy and as effective drug delivery means, but the fulfilment of these expectations requires overcoming at least two bottlenecks relative to their purification, namely the finalization of reliable and affordable protocols for: (i) EV sub-population selective isolation and (ii) the scalability of their production/isolation from complex biological fluids. In this work, we demonstrated that these objectives can be achieved by a conceptually new affinity chromatography platform composed of a macroporous epoxy monolith matrix functionalized with anti-CD63 nanobodies with afflux of samples and buffers regulated through a pump. Such a system successfully captured and released integral EVs from urine samples and showed negligible unspecific binding for circulating proteins. Additionally, size discrimination of eluted EVs was achieved by different elution approaches (competitive versus pH-dependent). The physical characteristics of monolith material and the inexpensive production of recombinant nanobodies make scaling-up the capture unit feasible and affordable. Additionally, the availability of nanobodies for further specific EV biomarkers will allow for the preparation of monolithic affinity filters selective for different EV subclasses.
Keywords: monolith chromatography; nanobodies; extracellular vesicles; affinity purification; CD63 monolith chromatography; nanobodies; extracellular vesicles; affinity purification; CD63

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MDPI and ACS Style

Neumair, J.; D’Ercole, C.; De March, M.; Elsner, M.; Seidel, M.; de Marco, A. Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine. Int. J. Mol. Sci. 2023, 24, 6131. https://doi.org/10.3390/ijms24076131

AMA Style

Neumair J, D’Ercole C, De March M, Elsner M, Seidel M, de Marco A. Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine. International Journal of Molecular Sciences. 2023; 24(7):6131. https://doi.org/10.3390/ijms24076131

Chicago/Turabian Style

Neumair, Julia, Claudia D’Ercole, Matteo De March, Martin Elsner, Michael Seidel, and Ario de Marco. 2023. "Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine" International Journal of Molecular Sciences 24, no. 7: 6131. https://doi.org/10.3390/ijms24076131

APA Style

Neumair, J., D’Ercole, C., De March, M., Elsner, M., Seidel, M., & de Marco, A. (2023). Macroporous Epoxy-Based Monoliths Functionalized with Anti-CD63 Nanobodies for Effective Isolation of Extracellular Vesicles in Urine. International Journal of Molecular Sciences, 24(7), 6131. https://doi.org/10.3390/ijms24076131

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