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. 2018 Nov 8;8:16550. doi: 10.1038/s41598-018-34845-2

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Isolation of human iPSC-derived CE lineage cells by CD200-negative sorting. (a) Flow cytometric analyses of SSEA-4, ITGB4, and CD200 (upper panels) or TRA-1-60 (lower panels) expression by differentiated iPSCs at weeks 10–15 of differentiation. ITGB4⁺/SSEA-4⁺ cells were considered to be CE lineage cells obtained from CD200- or TRA-1-60-negative populations, respectively. (b) Collection rate of iPSC-derived CE lineage cell population (ITGB4⁺/SSEA-4⁺) obtained by negative sorting procedures. Error bars, SD (n = 6). (c) Expression of corneal and non-target cell-related genes by the isolated iPSC-derived CE-like cells. Relative expression levels were set as 1 in TRA-1-60-negative samples (TRA). Error bars, SD (n = 9); *p < 0.05. (d) Representative phase-contrast images showing the cultivated iPSC-derived CE-like cells isolated by CD200- (upper panels) or TRA-1-60-negative sorting (lower panels). Right: magnified areas shown in the left panels. Asterisks: non-target colonies. Scale bars: 2 mm (left), 500 μm (right). (e) Numbers of non-target colonies in the cultivated iPSC-derived CE-like cells. Error bars, SD (n = 4); *p < 0.05.