Volume 156, Issue 1, Pages (January 2019)

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Volume 156, Issue 1, Pages 20-23 (January 2019) An Efficient Method for Cloning Gastrointestinal Stem Cells From Patients via Endoscopic Biopsies Marcin Duleba, Yutao Qi, Rajasekaran Mahalingam, Audrey-Ann Liew, Rahul Neupane, Kevin Flynn, Fabrizio Rinaldi, Matthew Vincent, Christopher P. Crum, Khek Yu Ho, Jason K. Hou, Jeffrey S. Hyams, Francisco A. Sylvester, Frank McKeon, Wa Xian Gastroenterology Volume 156, Issue 1, Pages 20-23 (January 2019) DOI: 10.1053/j.gastro.2018.08.062 Copyright © 2019 Terms and Conditions

Figure 1 Clonal analysis of colonic stem cells from endoscopic biopsies. A, Workflow of generating “libraries” of single cell derived colonies and subsequently 3-dimensional intestinal epithelium from 1-mm endoscopic biopsies. White light imaging of a typical endoscopic biopsy, representative images of 100–300 colonies derived from a typical biopsy, top view of in vitro intestinal epithelium generated from these stem cells differentiated in an air–liquid interface setting. Scale bar, 1000 μm. B, Individual colonies are sampled from the pool and grown in isolation as separate lines. C, Histologic analysis of in vitro-differentiated colonic epithelium via hematoxylin eosin staining, and immunofluorescence of antibodies to secretory cell markers Mucin 2, Chromogranin A, and Defensin alpha 6. Scale bar, 50 μm. Gastroenterology 2019 156, 20-23DOI: (10.1053/j.gastro.2018.08.062) Copyright © 2019 Terms and Conditions

Figure 2 Immortality and rapid expansion of colonic stem cells in vitro. A, Clonogenicity of single GFP-labeled colonic stem cell sorted to individual wells. B, Clonogenicity assay revealing nearly unchanged number of Rhodamine red-stained colonies grown 10 days after seeding of 2000 passaged colonic stem cells. C, Rapid expansion of a single cell to 1 billion cells in approximately 60 days. Gastroenterology 2019 156, 20-23DOI: (10.1053/j.gastro.2018.08.062) Copyright © 2019 Terms and Conditions