Abstract
Due to their high biological complexity, e.g., their close cell-to-cell contacts, cryopreservation of human pluripotent stem cells with standard slow-rate protocols often is inefficient and can hardly be standardized. Vitrification that means ultrafast freezing already showed very good viability and recovery rates for this sensitive cell system, but is only applicable for low cell numbers, bears a high risk of contamination, and can hardly be implemented under GxP regulations. In this chapter, a sterile plate-based vitrification method for adherent pluripotent stem cells and their derivatives is presented based on a procedure and device for human embryonic stem cells developed by Beier et al. (Cryobiology 66:8–16, 2013). This protocol overcomes the limitations of conventional vitrification procedures resulting in the highly efficient preservation of ready-to-use adherent pluripotent stem cells with the possibility of vitrifying cells in multi-well formats for direct application in high-throughput screenings.
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Acknowledgment
The work with human embryonic stem cells was permitted by the Robert Koch Institute (18. and 44. permission) and carried out according to German law.
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Neubauer, J.C., Stracke, F., Zimmermann, H. (2017). Chapter 17 Sterile Plate-Based Vitrification of Adherent Human Pluripotent Stem Cells and Their Derivatives Using the TWIST Method. In: Nagy, Z., Varghese, A., Agarwal, A. (eds) Cryopreservation of Mammalian Gametes and Embryos. Methods in Molecular Biology, vol 1568. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6828-2_17
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DOI: https://doi.org/10.1007/978-1-4939-6828-2_17
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