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A qPCR ScoreCard quantifies the differentiation potential of human pluripotent stem cells

Nature Biotechnology volume 33, pages 11821192 (2015) | Download Citation

Abstract

Research on human pluripotent stem cells has been hampered by the lack of a standardized, quantitative, scalable assay of pluripotency. We previously described an assay called ScoreCard that used gene expression signatures to quantify differentiation efficiency. Here we report an improved version of the assay based on qPCR that enables faster, more quantitative assessment of functional pluripotency. We provide an in-depth characterization of the revised signature panel (commercially available as the TaqMan hPSC Scorecard Assay) through embryoid body and directed differentiation experiments as well as a detailed comparison to the teratoma assay. We further show that the improved ScoreCard enables a wider range of applications, such as screening of small molecules, genetic perturbations and assessment of culture conditions. Our approach can be extended beyond stem cell applications to characterize and assess the utility of other cell types and lineages.

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Acknowledgements

We would like to thank all members of the Meissner laboratory as well as D. Tieberg and U. Lakshmipathy for their valuable support and insightful feedback, L. Solomon and L. Gaffney for graphical support, I.S. Kim and B. Bernstein for the JQ1 inhibitor molecule and insight, L. Rubin for the partially reprogrammed cell lines and C. MacGillivray (HSCI Histology core) for teratoma sectioning, fixation and histochemical staining for markers of the three germ layers. A.M.T. is supported by the NIH under Ruth L. Kirschstein National Research Service Award (NRSA) fellowship 5F32DK095537. This work was supported by the National Institute of General Medical Sciences (NIGMS) (P01GM099117), National Human Genome Research Institute (NHGRI) (1P50HG006193), the New York Stem Cell Foundation, and a research grant from Life Technologies. A.M. is a New York Stem Cell Foundation, Robertson Investigator.

Author information

Affiliations

  1. Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

    • Alexander M Tsankov
    • , Veronika Akopian
    • , Ramona Pop
    • , Casey A Gifford
    •  & Alexander Meissner
  2. Harvard Stem Cell Institute, Cambridge, Massachusetts, USA.

    • Alexander M Tsankov
    • , Veronika Akopian
    • , Ramona Pop
    • , Sundari Chetty
    • , Casey A Gifford
    • , Laurence Daheron
    •  & Alexander Meissner
  3. Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts, USA.

    • Alexander M Tsankov
    • , Veronika Akopian
    • , Ramona Pop
    • , Sundari Chetty
    • , Casey A Gifford
    •  & Alexander Meissner
  4. Department of Pathology and Cell Biology, Columbia University Medical Center, New York, New York, USA.

    • Nadejda M Tsankova
  5. Department of Pathology, Icahn School of Medicine at Mount Sinai, New York, New York, USA.

    • Nadejda M Tsankova
  6. Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, New York, USA.

    • Nadejda M Tsankova

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Contributions

A.M.T. and A.M. conceived and designed the study. A.M.T. optimized all experiments and performed all analysis, V.A. performed cell culture and ran qPCR plates, R.P. performed FACS analysis and ran qPCR plates, C.A.G. trained A.M.T. in cell culture and provided experimental advice, S.C. performed PiPSC embryoid body experiments with advice from L.D., N.M.T. performed all pathological and histological annotations, A.M.T. and A.M. interpreted the data and wrote the manuscript.

Competing interests

Harvard University has filed a patent on the original ScoreCard technology and a licensing agreement with Life Technologies (now part of Thermo Fisher). A.M. is an advisor to Life Technologies.

Corresponding author

Correspondence to Alexander Meissner.

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DOI

https://doi.org/10.1038/nbt.3387

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