Abstract
This chapter describes a detailed protocol on human pluripotent stem cells (hPSCs) cultivation as matrix-free cell-only aggregates in defined and xeno-free culture medium in stirred tank bioreactors (STBRs). Starting with a frozen stock pre-expanded on conventional culture dishes (2D), the ultimate process is performed in 150 mL culture scale in stirred tank bioreactors (3D) and is designed to produce up to 500 million pluripotent hPSC within 7 days. The culture strategy includes perfusion-based cell feeding facilitating process control, automation, and higher cell yields. Ultimately, this detailed protocol describes an important step for generating a defined starting cell population for directed lineage differentiation and subsequently fueling human cell-based assays and regenerative medicine approaches.
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References
Inoue H, Nagata N, Kurokawa H, Yamanaka S (2014) iPS cells: a game changer for future medicine. EMBO J 33:409–417
Kempf H, Andree B, Zweigerdt R (2016) Largescale production of human pluripotent stem cell derived cardiomyocytes. Adv Drug Deliv Rev 96:18–30
Serra M, Brito C, Correia C, Alves PM (2012) Process engineering of human pluripotent stem cells for clinical application. Trends Biotechnol 30:350–359
Zweigerdt R (2009) Large scale production of stem cells and their derivatives. Adv Biochem Eng Biotechnol 114:201–235
Serra M, Brito C, Sousa MF, Jensen J, Tostões R, Clemente J et al (2010) Improving expansion of pluripotent human embryonic stem cells in perfused bioreactors through oxygen control. J Biotechnol 148:208–215
Kropp C, Kempf H, Halloin C, Robles-Diaz D, Franke A, Scheper T et al (2016) Impact of feeding strategies on the scalable expansion of human pluripotent stem cells in single-use stirred tank bioreactors. Stem Cells Transl Med 5:1289–1301
Kempf H, Olmer R, Kropp C, Ruckert M, Jara-Avaca M, Robles-Diaz D et al (2014) Controlling expansion and cardiomyogenic differentiation of human pluripotent stem cells in scalable suspension culture. Stem Cell Reports 6:1132–1146
Olmer R, Engels L, Usman A, Menke S, Malik MNH, Pessler F et al (2018) Differentiation of human pluripotent stem cells into functional endothelial cells in scalable suspension culture. Stem Cell Reports 5:1657–1672
Chen G, Gulbranson DR, Hou Z, Bolin JM, Ruotti V, Probasco MD et al (2011) Chemically defined conditions for human iPSC derivation and culture. Nat Methods 8:424–429
Acknowledgments
This work was supported by StemBANCC (support from the Innovative Medicines Initiative joint undertaking under grant 115439-2, whose resources are composed of financial contribution from the European Union [FP7/2007-2013] and EFPIA companies’ in-kind contribution). RZ received funding from the German Research Foundation (DFG; including the Cluster of Excellence REBIRTH EXC 62/2 and grants ZW64/4-1, KFO311, and ZW64/7-1), the German Ministry for Education and Science (BMBF; grants: 13 N14086, 01EK1601A, and 01EK1602A) and by the European Union (TECHNOBEAT; grant 66724).
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Manstein, F., Halloin, C., Zweigerdt, R. (2019). Human Pluripotent Stem Cell Expansion in Stirred Tank Bioreactors. In: Mandenius, CF., Ross, J. (eds) Cell-Based Assays Using iPSCs for Drug Development and Testing. Methods in Molecular Biology, vol 1994. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9477-9_7
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DOI: https://doi.org/10.1007/978-1-4939-9477-9_7
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Online ISBN: 978-1-4939-9477-9
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