292 Chapter 11 been described[35]. Here, we found the PSC Cryopreservation Kit is the most optimal condition as compared to 3 others (data not shown) and used this medium for the efficient freezing of spheroids. After cryopreservation, viability remains high (Figures 4B and Figure 4C), but CSs’ electrophysiological properties are affected and a period of incubation after thawing is required. Indeed, one week after thawing, CSs displayed spontaneous beating activity and calcium handling. However, it has been described that fresh and recovered hiPSC-CMs do not always show identical molecular and physiological properties[36]. This limitation needs to be considered when cryopreserved hiPSC-CMs are used for assessing drug-induced cardiac read-outs. Overall, we propose a step-by-step protocol to efficiently generate CSs which are suitable for downstream applications such as disease modeling and HT drug screening. Disclosures The authors have nothing to disclose. Acknowledgments We would like to acknowledge VALA sciences for the Cyteseer software package and optimization of the automated 3D calcium analysis. We wish to acknowledge grant support from the PLN foundation (RM). P.A.D. and F.S. are supported by CUREPLaN Leducq. J.P.G.S. is supported by H2020-EVICARE (#725229) of the European Research Council (ERC). J.W.B. is supported by the UMC Utrecht Clinical Fellowship, Netherlands Heart Institute Fellowship, and CVON-Dosis young talent grant; Netherlands Heart Foundation (CVON-Dosis 2014–40). NC is supported by the Gravitation Program “Materials Driven Regeneration” by the Netherlands Organization for Scientific Research (RegmedXB #024.003.013), and the Marie SkłodowskaCurie Actions (Grant agreement RESCUE #801540). V. S.-P. is supported by the Alliance Fund (UMCU, UU, TU/e). A.v.M. is supported by the EU-funded project BRAVE (H2020, ID:874827)
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