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Chemically defined conditions for human iPSC derivation and culture

Nature Methods volume 8, pages 424–429 (2011)Cite this article

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Abstract

We re-examine the individual components for human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) culture and formulate a cell culture system in which all protein reagents for liquid media, attachment surfaces and splitting are chemically defined. A major improvement is the lack of a serum albumin component, as variations in either animal- or human-sourced albumin batches have previously plagued human ESC and iPSC culture with inconsistencies. Using this new medium (E8) and vitronectin-coated surfaces, we demonstrate improved derivation efficiencies of vector-free human iPSCs with an episomal approach. This simplified E8 medium should facilitate both the research use and clinical applications of human ESCs and iPSCs and their derivatives, and should be applicable to other reprogramming methods.

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Figure 1: Albumin is not required for human ESC culture.
Figure 2: Essential medium components for human ESC survival and proliferation.
Figure 3: Vitronectin-coated surfaces support human ESCs and iPSCs cultured in E8.
Figure 4: Reprogramming fibroblast cells in fully defined conditions.
Figure 5: Derivation of human iPSCs directly from biopsy samples in chemically defined conditions.

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Acknowledgements

This work was supported by The Charlotte Geyer Foundation, the Morgridge Institute for Research, US National Institutes of Health grant UO1ES017166 (to J.A.T.) and National Institutes of Health contract RR-05-19 (to J.A.T.). We thank K. Eastman for editorial assistance, and C. Dewey, R. Stewart and A. Elwell for their assistance with gene expression analysis.

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Authors and Affiliations

  1. Morgridge Institute for Research, Madison, Wisconsin, USA

    Guokai Chen, Daniel R Gulbranson, Zhonggang Hou, Jennifer M Bolin, Victor Ruotti, Mitchell D Probasco, Sara E Howden, Nicole R Diol, Nicholas E Propson, Ryan Wagner, Garrett O Lee, Jessica Antosiewicz-Bourget & James A Thomson

  2. Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, USA

    Guokai Chen, Daniel R Gulbranson, Zhonggang Hou, Sara E Howden, Nicole R Diol, Nicholas E Propson, Ryan Wagner, Garrett O Lee & James A Thomson

  3. The Genome Center of Wisconsin, University of Wisconsin, Madison, Wisconsin, USA

    Guokai Chen, Daniel R Gulbranson, Zhonggang Hou, Sara E Howden, Nicole R Diol, Nicholas E Propson, Ryan Wagner, Garrett O Lee & James A Thomson

  4. Wisconsin National Primate Research Center, Madison, Wisconsin, USA

    Kimberly Smuga-Otto

  5. Dermatology Department, University of Wisconsin, Madison, Wisconsin, USA

    Joyce M C Teng

  6. Department of Molecular, Cellular and Developmental Biology, University of California Santa Barbara, Santa Barbara, California, USA

    James A Thomson

Authors
  1. Guokai Chen
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Contributions

G.C. and J.A.T. conceived the experiment; G.C., D.R.G., J.M.B., K.S.-O. and S.E.H. performed the reprogramming; Z.H., G.C. and N.E.P. produced vitronectin; G.C., D.R.G., J.M.B., N.R.D., G.O.L. and J.A.-B. performed the cell culture test; G.C., M.D.P. and R.W. derived fibroblasts; J.M.C.T. obtained the skin biopsy; V.R. and G.C. analyzed global expression; and G.C. and J.A.T. wrote the paper.

Corresponding author

Correspondence to James A Thomson.

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Competing interests

J.A.T. is a founder, stockowner, consultant and board member of Cellular Dynamics International (CDI), and serves as scientific advisor to and has financial interests in Tactics II Stem Cell Ventures.

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Supplementary Figures 1–4 and Supplementary Tables 1–3 (PDF 6386 kb)

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Chen, G., Gulbranson, D., Hou, Z. et al. Chemically defined conditions for human iPSC derivation and culture. Nat Methods 8, 424–429 (2011). https://doi.org/10.1038/nmeth.1593

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