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Dynamic suspension culture for scalable expansion of undifferentiated human pluripotent stem cells

Nature Protocols volume 6, pages 572–579 (2011)Cite this article

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Abstract

Human pluripotent (embryonic or induced) stem cells (hPSCs) have many potential applications, not only for research purposes but also for clinical and industrial uses. While culturing these cells as undifferentiated lines, an adherent cell culture based on supportive layers or matrices is most often used. However, the use of hPSCs for industrial or clinical applications requires a scalable, reproducible and controlled process. Here we present a suspension culture system for undifferentiated hPSCs, based on a serum-free medium supplemented with interleukins and basic fibroblast growth factor, suitable for the mass production of these cells. The described system supports a suspension culture of hPSC lines, in both static and dynamic cultures. Results showed that hPSCs cultured with the described dynamic method maintained all hPSC features after 20 passages, including stable karyotype and pluripotency, and increased in cell numbers by 25-fold in 10 d. Thus, the described suspension method is suitable for large-scale culture of undifferentiated hPSCs.

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Figure 1: Sphere morphology.
Figure 2: Teratoma formation.

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Acknowledgements

The authors thank M. Revel of Weizmann Institute of Science for providing the IL6IL6-receptor chimera. This research was supported by the Technion Research and Development Foundation (TRDF). J.I.-E. holds the Sylvia and Stanley Shirvan Chair in Cell and Tissue Regeneration Research at the Technion-Israel Institute of Technology.

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Authors and Affiliations

  1. The Sohnis and Forman Families Stem Cell Center, the Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel

    Michal Amit, Ilana Laevsky, Yael Miropolsky, Kohava Shariki, Meital Peri & Joseph Itskovitz-Eldor

  2. Department of Obstetrics and Gynecology, Rambam Health Care Campus, Haifa, Israel

    Joseph Itskovitz-Eldor

Authors
  1. Michal Amit
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  2. Ilana Laevsky
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  3. Yael Miropolsky
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  4. Kohava Shariki
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  5. Meital Peri
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  6. Joseph Itskovitz-Eldor
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Contributions

M.A. developed the concept, designed the experiments, performed data analysis and wrote the manuscript; I.L., Y.M., K.S. and M.P. provided technical assistance. J.I.-E. developed the concept, provided overall guidance, performed data analysis and interpretation, revised and provided final approval of the manuscript.

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Correspondence to Joseph Itskovitz-Eldor.

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The authors declare no competing financial interests.

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Amit, M., Laevsky, I., Miropolsky, Y. et al. Dynamic suspension culture for scalable expansion of undifferentiated human pluripotent stem cells. Nat Protoc 6, 572–579 (2011). https://doi.org/10.1038/nprot.2011.325

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