Embryonic Stem Cells/Induced Pluripotent Stem Cells
Identification of Unsafe Human Induced Pluripotent Stem Cell Lines Using a Robust Surrogate Assay for Pluripotency†
Version of Record online: 23 AUG 2013
Copyright © 2013 AlphaMed Press
Volume 31, Issue 8, pages 1498–1510, August 2013
How to Cite
Polanco, J. C., Ho, M. S.H., Wang, B., Zhou, Q., Wolvetang, E., Mason, E., Wells, C. A., Kolle, G., Grimmond, S. M., Bertoncello, I., O'Brien, C. and Laslett, A. L. (2013), Identification of Unsafe Human Induced Pluripotent Stem Cell Lines Using a Robust Surrogate Assay for Pluripotency. STEM CELLS, 31: 1498–1510. doi: 10.1002/stem.1425
Author contributions: J.C.P.: conception and design, collection and/or assembly of data, data analysis and interpretation, and manuscript writing; M.S.H.H.: conception and design and collection and/or assembly of data; B.W., Q.Z., and G.K.: collection and/or assembly of data; E.W.: provision of study material; E.M.: data analysis and interpretation; C.A.W.: data analysis and interpretation and manuscript writing; S.M.G.: collection and/or assembly of data and data analysis and interpretation; I.B.: conception and design; C.O.B.: conception and design, manuscript writing, and data analysis and interpretation; A.L.L.: conception and design, collection and/or assembly of data, data analysis and interpretation, provision of study material, manuscript writing, financial support, and final approval of manuscript.
- Issue online: 23 AUG 2013
- Version of Record online: 23 AUG 2013
- Accepted manuscript online: 3 JUN 2013 01:35AM EST
- Manuscript Accepted: 14 APR 2013
- Manuscript Received: 21 FEB 2013
- Australian Stem Cell Centre
- New South Wales and Victorian Government Stem Cell Research Grant Program
- Partner Investigator on the Australian Research Council (ARC)
- Special Research Initiative in Stem Cell Science, Stem Cells Australia
Additional Supporting Information may be found in the online version of this article.
|STEM_1425_sm_SuppFigure1.pdf||265K||Supplementary Figure S1. Principal Component Analysis (PCA). PCA generated using quantile normalised data from the arrays conducted in this study on the hiPSC lines [L]IMR90C2 and [L]ForeskinC1 and from the hESC lines H9 and MEL1 from Kolle et al., 2009. [A] Separation of hiPSC and hESC samples on component 1 is likely explained by differences in the array version between the published hESC data (V2) and newer hiPSC experiments (V3) and component 2 appears to be driven by TG30/GCTM-2 fractionation. [B] Absolute measure of the percentage variance of each principal component indicating that the data can be explained by the first three components.|
|STEM_1425_sm_SuppFigure2.pdf||216K||Supplementary Figure S2. Persistence of P7 cells does not cease with long term in-vitro passaging and presence of OCT4 in P4 cultures. (A): Comparative analysis of Colony Forming Assays (CFA) between late-passage (<50 passages) lentivirally-derived [L]hIPSForeskinC1 and hESC-MEL1 control line. A defined number of P4 (TG30Neg-GCTM-2Neg) cells were cultured post-FACS in 12-well plates, generating an elevated number of hPSC-like colonies only for the lentiviral hiPSC samples. Data collected from three independent CFA (n=3), each assayed in triplicate (mean ± SEM). (B): Immunofluorescence for pluripotency markers. Colonies generated by P4 (TG30Neg-GCTM-2Neg) cells from late-passage lentiviral hiPSC samples, exhibited immunoreactivity to the pluripotency markers OCT4 (red) and NANOG (green). (C): Quantification of OCT4 protein in the P4 (TG30Neg-GCTM-2Neg) population from both hiPSC and hESC lines. P4 exhibits a low level of OCT4 protein and requires the highly sensitive TaqManR Protein Assays for quantification. A defined number of P4 (TG30Neg-GCTM-2Neg) cells was collected by FACS from standard cultures of hiPSC lines (lentiviral [L]hIPS-IMR90C2 and [L]hIPS-ForeskinC1, retroviral [R]hIPS-PDL-D1C6) and hESC control (MEL1), followed by OCT4 protein quantification. Expression levels are shown relative to hESC-MEL1 set to 100%. Data from three independent experiments (n=3), assayed in triplicates, error bars represent SEM. Asterisks highlight statistical significance with respect to the control sample hESC-MEL1. *P<0.05, **P<0.01, ***P<0.001; ns, P<0.05, with a 95% confidence interval.|
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