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. 2018 Oct 18;8:15410. doi: 10.1038/s41598-018-33747-7

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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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Spheroid formation assay to identify GI radioprotectors. Schematic representation illustrating how stem cell-enriched epithelial spheroid cultures are established (panel a). Schematic outlining how the spheroid formation assay (SFA) is performed (see text for details, panel b). Z-stack images of treated stem cell-enriched epithelial spheroid cultures are stitched together to visualize spheroids (panel c). A representative image of the stitched z-stack is shown (panel d). Stem cell-enriched epithelial spheroid cultures treated with vehicle (PBS) or 2 mM WR-1065 for 2 h were exposed to the indicated doses of ionizing radiation. Spheroids were immediately dissociated into single cells, replated in Matrigel and imaged 5 days later (panel e) (Scale bars, 1000 μm). Spheroids larger than 150 μm in diameter were quantified and mean surviving fraction is plotted. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 by two-tailed, Student’s t test (N = 3 per group). Error bars are ± SEM.