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Fabrication of synthetic polymer coatings and their use in feeder-free culture of human embryonic stem cells

Nature Protocols volume 6, pages 1037–1043 (2011)Cite this article

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Abstract

The culture of human embryonic stem (hES) cells in defined and xenogeneic-free conditions will contribute substantially to future biotechnological and medical applications. To achieve this goal, we developed the first fully defined synthetic polymer coating poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl)ammonium hydroxide] (PMEDSAH) that sustains long-term growth of hES cells in different culture media. Here we describe a detailed protocol for the reproducible fabrication of PMEDSAH coating on tissue culture polystyrene dishes, and for the feeder-free culture of hES cells on PMEDSAH coating in defined culture medium. This culture system represents a key step toward the fully defined and xenogeneic-free culture of hES cells.

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Figure 1: Reactor setup used for surface-initiated graft polymerization of PMEDSAH on TCPS dishes.
Figure 2: Culture of hES cells on PMEDSAH-coated TCPS dishes.

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Acknowledgements

H.N. acknowledges funding from the University of Michigan Rackham Predoctoral Fellowship. J.L. gratefully acknowledges support from the NSF in form of a CAREER grant and funding from the NSF under the MRI program. This research was supported by NIH grants P20 GM-069985, R01 DE016530 and the NIDCR T32 Tissue Engineering and Regeneration Training Program.

Author information

Author notes

  1. Himabindu Nandivada

    Present address: Present address: BD Biosciences–Discovery Labware, Bedford, Massachusetts, USA.,

  2. Himabindu Nandivada and Luis G Villa-Diaz: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Chemical Engineering, University of Michigan, Ann Arbor, Michigan, USA

    Himabindu Nandivada & Joerg Lahann

  2. Department of Biologic and Materials Sciences, University of Michigan, Ann Arbor, Michigan, USA

    Luis G Villa-Diaz & Paul H Krebsbach

  3. Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, Michigan, USA

    K Sue O'Shea

  4. Department of Obstetrics and Gynecology, University of Michigan, Ann Arbor, Michigan, USA

    Gary D Smith

  5. Department of Urology, University of Michigan, Ann Arbor, Michigan, USA

    Gary D Smith

  6. Department of Molecular and Integrated Physiology, University of Michigan, Ann Arbor, Michigan, USA

    Gary D Smith

  7. Department of Reproductive Sciences Program, University of Michigan, Ann Arbor, Michigan, USA

    Gary D Smith

  8. Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan, USA

    Paul H Krebsbach

  9. Department of Materials Science and Engineering, University of Michigan, Ann Arbor, Michigan, USA

    Joerg Lahann

  10. Department of Macromolecular Science and Engineering, University of Michigan, Ann Arbor, Michigan, USA

    Joerg Lahann

Authors
  1. Himabindu Nandivada
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  2. Luis G Villa-Diaz
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  5. Paul H Krebsbach
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Contributions

This protocol was developed and written by H.N. and L.G.V.-D. after obtaining the material from all authors. H.N. designed the 'Fabrication of PMEDSAH-coated TCPS dishes' portion of the protocol; L.G.V.-D. designed the 'Establishment of hES cell cultures on PMEDSAH-coated TCPS dishes' portion of the protocol. K.S.O., G.D.S., P.H.K. and J.L. supervised the project.

Corresponding authors

Correspondence to Luis G Villa-Diaz, Gary D Smith or Joerg Lahann.

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The authors declare no competing financial interests.

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Nandivada, H., Villa-Diaz, L., O'Shea, K. et al. Fabrication of synthetic polymer coatings and their use in feeder-free culture of human embryonic stem cells. Nat Protoc 6, 1037–1043 (2011). https://doi.org/10.1038/nprot.2011.342

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