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Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells

Achieving good cell recovery after cryopreservation is an essential process when working with induced pluripotent stem cells (iPSC). Optimized freezing and thawing methods are required for good cell attachment and survival. In this review, we concentrate on these two aspects, freezing and thawing, b...

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Autores principales: Uhrig, Markus, Ezquer, Fernando, Ezquer, Marcelo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8909336/
https://www.ncbi.nlm.nih.gov/pubmed/35269421
http://dx.doi.org/10.3390/cells11050799
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author Uhrig, Markus
Ezquer, Fernando
Ezquer, Marcelo
author_facet Uhrig, Markus
Ezquer, Fernando
Ezquer, Marcelo
author_sort Uhrig, Markus
collection PubMed
description Achieving good cell recovery after cryopreservation is an essential process when working with induced pluripotent stem cells (iPSC). Optimized freezing and thawing methods are required for good cell attachment and survival. In this review, we concentrate on these two aspects, freezing and thawing, but also discuss further factors influencing cell recovery such as cell storage and transport. Whenever a problem occurs during the thawing process of iPSC, it is initially not clear what it is caused by, because there are many factors involved that can contribute to insufficient cell recovery. Thawing problems can usually be solved more quickly when a certain order of steps to be taken is followed. Under optimized conditions, iPSC should be ready for further experiments approximately 4–7 days after thawing and seeding. However, if the freezing and thawing protocols are not optimized, this time can increase up to 2–3 weeks, complicating any further experiments. Here, we suggest optimization steps and troubleshooting options for the freezing, thawing, and seeding of iPSC on feeder-free, Matrigel™-coated, cell culture plates whenever iPSC cannot be recovered in sufficient quality. This review applies to two-dimensional (2D) monolayer cell culture and to iPSC, passaged, frozen, and thawed as cell aggregates (clumps). Furthermore, we discuss usually less well-described factors such as the cell growth phase before freezing and the prevention of osmotic shock during thawing.
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spelling pubmed-89093362022-03-11 Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells Uhrig, Markus Ezquer, Fernando Ezquer, Marcelo Cells Review Achieving good cell recovery after cryopreservation is an essential process when working with induced pluripotent stem cells (iPSC). Optimized freezing and thawing methods are required for good cell attachment and survival. In this review, we concentrate on these two aspects, freezing and thawing, but also discuss further factors influencing cell recovery such as cell storage and transport. Whenever a problem occurs during the thawing process of iPSC, it is initially not clear what it is caused by, because there are many factors involved that can contribute to insufficient cell recovery. Thawing problems can usually be solved more quickly when a certain order of steps to be taken is followed. Under optimized conditions, iPSC should be ready for further experiments approximately 4–7 days after thawing and seeding. However, if the freezing and thawing protocols are not optimized, this time can increase up to 2–3 weeks, complicating any further experiments. Here, we suggest optimization steps and troubleshooting options for the freezing, thawing, and seeding of iPSC on feeder-free, Matrigel™-coated, cell culture plates whenever iPSC cannot be recovered in sufficient quality. This review applies to two-dimensional (2D) monolayer cell culture and to iPSC, passaged, frozen, and thawed as cell aggregates (clumps). Furthermore, we discuss usually less well-described factors such as the cell growth phase before freezing and the prevention of osmotic shock during thawing. MDPI 2022-02-24 /pmc/articles/PMC8909336/ /pubmed/35269421 http://dx.doi.org/10.3390/cells11050799 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Review
Uhrig, Markus
Ezquer, Fernando
Ezquer, Marcelo
Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title_full Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title_fullStr Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title_full_unstemmed Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title_short Improving Cell Recovery: Freezing and Thawing Optimization of Induced Pluripotent Stem Cells
title_sort improving cell recovery: freezing and thawing optimization of induced pluripotent stem cells
topic Review
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8909336/
https://www.ncbi.nlm.nih.gov/pubmed/35269421
http://dx.doi.org/10.3390/cells11050799
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