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Extracellular antifreeze protein significantly enhances the cryopreservation of cell monolayers

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Tomás, Ruben M. F., Bailey, Trisha L., Hasan, Muhammad and Gibson, Matthew I. (2019) Extracellular antifreeze protein significantly enhances the cryopreservation of cell monolayers. Biomacromolecules, 20 (10). pp. 3864-3872. doi:10.1021/acs.biomac.9b00951

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Official URL: https://doi.org/10.1021/acs.biomac.9b00951

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Abstract

The cryopreservation of cells underpins many areas of biotechnology, healthcare, and fundamental science by enabling the banking and distribution of cells. Cryoprotectants are essential to prevent cold-induced damage. Here, we demonstrate that extracellular localization of antifreeze proteins can significantly enhance post-thaw recovery of mammalian cell monolayers cryopreserved using dimethyl sulfoxide, whereas they show less benefit in suspension cryopreservation. A type III antifreeze protein (AFPIII) was used as the macromolecular ice recrystallization inhibitor and its intra/extracellular locations were controlled by using Pep-1, a cell-penetrating peptide. Flow cytometry and confocal microscopy confirmed successful delivery of AFPIII. The presence of extracellular AFPIII dramatically increased post-thaw recovery in a challenging 2-D cell monolayer system using just 0.8 mg·mL–1, from 25% to over 60%, whereas intracellularly delivered AFPIII showed less benefit. Interestingly, the antifreeze protein was less effective when used in suspension cryopreservation of the same cells, suggesting that the cryopreservation format is also crucial. These observations show that, in the discovery of macromolecular cryoprotectants, intracellular delivery of ice recrystallization inhibitors may not be a significant requirement under “slow freezing” conditions, which will help guide the design of new biomaterials, in particular, for cell storage.

Item Type: Journal Article
Subjects: Q Science > QH Natural history > QH301 Biology
Divisions: Faculty of Science > Chemistry
SWORD Depositor: Library Publications Router
Library of Congress Subject Headings (LCSH): Cryopreservation of organs, tissues, etc -- Methods, Cells -- Preservation, Biotechnology, Freeze-drying -- Methods
Journal or Publication Title: Biomacromolecules
Publisher: American Chemical Society (ACS)
ISSN: 1526-4602
Official Date: 14 October 2019
Dates:
DateEvent
14 October 2019Published
9 September 2019Available
9 September 2019Accepted
Date of first compliant deposit: 12 November 2019
Volume: 20
Number: 10
Page Range: pp. 3864-3872
DOI: 10.1021/acs.biomac.9b00951
Status: Peer Reviewed
Publication Status: Published
Access rights to Published version: Open Access
RIOXX Funder/Project Grant:
Project/Grant IDRIOXX Funder NameFunder ID
CRYOMAT 638661H2020 European Research Councilhttp://dx.doi.org/10.13039/100010663
EP/L015307/1RT[EPSRC] Engineering and Physical Sciences Research Councilhttp://dx.doi.org/10.13039/501100000266
BB/R506588/1[BBSRC] Biotechnology and Biological Sciences Research Councilhttp://dx.doi.org/10.13039/501100000268
BB/M01228X/1[BBSRC] Biotechnology and Biological Sciences Research Councilhttp://dx.doi.org/10.13039/501100000268
789182H2020 European Research Councilhttp://dx.doi.org/10.13039/100010663
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