Misleading Pore Size Measurements in Gelatin and Alginate Hydrogels Revealed by Confocal Microscopy

Franco, Alexis;Van Durme, Bo;Van Vlierberghe, Sandra;Dupont, Christine
(2024) Tissue Engineering. Part C. Methods — Vol. 30, n° 7, p. 307-313 (2024)

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Authors
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  • Van Durme, BoPolymer Chemistry and Biomaterials Group, Centre of Macromolecular Chemistry, Ghent University, Ghent, Belgium
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  • Van Vlierberghe, SandraPolymer Chemistry and Biomaterials Group, Centre of Macromolecular Chemistry, Ghent University, Ghent, Belgium
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Abstract
It is a well-documented phenomenon that the porous structure of hydrogels observed with vacuum-based imaging techniques is generated during the freezing and drying process employed prior to observation. Nevertheless, vacuum-based techniques, such as scanning electron microscopy (SEM), are still being commonly used tomeasure pore sizes in hydrogels, which is often not representative of the actual pore size in hydrated conditions. The frequent underestimation of the impact of freezing and drying on hydrogel structures could stem from a lack of crossfertilization between materials science and biomedical or food science communities, or from the simplicity and visually appealing nature of SEM imaging, which may lead to an overemphasis on its use. Our study provides a straightforward and impactful way of pinpointing this phenomenon exploiting two hydrogels ubiquitously applied in tissue engineering, including gelatin methacryloyl and alginate as proof-of-concept hydrogels. By comparing images of the samples in the native hydrated state, followed by freezing, freeze-drying, and rehydration using SEM and confocal microscopy, we highlight discrepancies between hydrogel pore sizes in the hydrated versus the dry state. To conclude, our study offers recommendations for researchers seeking insight in hydrogel properties and emphasizes key factors that require careful control when using SEMas a characterization tool.
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Citations

Franco, A., Van Durme, B., Van Vlierberghe, S., & Dupont, C. (2024). Misleading Pore Size Measurements in Gelatin and Alginate Hydrogels Revealed by Confocal Microscopy. Tissue Engineering. Part C. Methods, 30(7), 307-313. https://doi.org/10.1089/ten.tec.2024.0117 (Original work published 2024)