Cellular and Molecular Bioengineering

, Volume 7, Issue 2, pp 172–183 | Cite as

Developing Defined and Scalable 3D Culture Systems for Culturing Human Pluripotent Stem Cells at High Densities

  • Yuguo Lei
  • Daeun Jeong
  • Jifang Xiao
  • David V. SchafferEmail author


Human pluripotent stem cells (hPSCs)—including embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs)—are very promising candidates for cell therapies, tissue engineering, high throughput pharmacology screens, and toxicity testing. These applications require large numbers of high quality cells; however, scalable production of human pluripotent stem cells and their derivatives at a high density and under well-defined conditions has been a challenge. We recently reported a simple, efficient, fully defined, scalable, and good manufacturing practice (GMP) compatible 3D culture system based on a thermoreversible hydrogel for hPSC expansion and differentiation. Here, we describe additional design rationale and characterization of this system. For instance, we have determined that culturing hPSCs as a suspension in a liquid medium can exhibit lower volumetric yields due to cell agglomeration and possible shear force-induced cell loss. By contrast, using hydrogels as 3D scaffolds for culturing hPSCs reduces aggregation and may insulate from shear forces. Additionally, hydrogel-based 3D culture systems can support efficient hPSC expansion and differentiation at a high density if compatible with hPSC biology. Finally, there are considerable opportunities for future development to further enhance hydrogel-based 3D culture systems for producing hPSCs and their progeny.


Human embryonic stem cells Induced pluripotent stem cells 3D culture system Thermoreversible hydrogel 



This work was supported by California Institute of Regenerative Medicine Grant RT2-02022, a California Institute for Regenerative Medicine training grant T1-00007 fellowship (to Y. L.), and NIH 1R01ES020903-01.

Conflict of interest

Yuguo Lei, Daeun Jeong, Jifang Xiao and David V. Schaffer declare that they have no conflicts of interest.

Ethical standings

All animal studies were carried out in accordance with NIH and University of California, Berkeley guidelines, and approved by the Animal Care and Use Committee of the University of California, Berkeley. No human studies were carried out by the authors for this article.


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Copyright information

© Biomedical Engineering Society 2014

Authors and Affiliations

  • Yuguo Lei
    • 1
    • 2
    • 4
    • 5
  • Daeun Jeong
    • 3
  • Jifang Xiao
    • 2
  • David V. Schaffer
    • 1
    • 2
    • 4
    • 5
    Email author
  1. 1.Department of Chemical and Biomolecular EngineeringUniversity of California, BerkeleyBerkeleyUSA
  2. 2.Department of BioengineeringUniversity of California, BerkeleyBerkeleyUSA
  3. 3.Department of Molecular and Cell BiologyUniversity of California, BerkeleyBerkeleyUSA
  4. 4.California Institute for Quantitative BiosciencesUniversity of California, BerkeleyBerkeleyUSA
  5. 5.Helen Wills Neuroscience InstituteUniversity of California, BerkeleyBerkeleyUSA

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