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Columbia University Irving Medical Center

3 ARTICLES PUBLISHED IN JoVE

Biology

Imaging of mtHyPer7, a Ratiometric Biosensor for Mitochondrial Peroxide, in Living Yeast Cells

Emily Jie-Ning Yang¹, Istvan R. Boldogh^1,2, Haojie Ji², Liza Pon^1,2, Theresa C. Swayne^1,2

¹Department of Pathology and Cell Biology, Columbia University Irving Medical Center, ²Confocal and Specialized Microscopy Shared Resource in the Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center

Hydrogen peroxide (H₂O₂) is both a source of oxidative damage and a signaling molecule. This protocol describes how to measure mitochondrial H₂O₂ using mitochondria-targeted HyPer7 (mtHyPer7), a genetically encoded ratiometric biosensor, in living yeast. It details how to optimize imaging conditions and perform quantitative cellular and subcellular analysis using freely available software.

Biology

Establishment and Characterization of Patient-derived Gastric Organoids from Biopsies of Benign Gastric Body and Antral Epithelium

Kole H. Buckley¹, Keely A. Beyries¹, Sandra Ryeom², Sam S. Yoon², Bryson W. Katona¹

¹Division of Gastroenterology and Hepatology, University of Pennsylvania Perelman School of Medicine, ²Department of Surgery, Columbia University Irving Medical Center

Gastric patient-derived organoids find increasing use in research, yet formal protocols for generating human gastric organoids from single-cell digests with standardized seeding density are lacking. This protocol presents a detailed method for reliably creating gastric organoids from biopsy tissue obtained during upper endoscopy.

Biology

CRISPR-Cas9 Mediated Gene Deletion in Human Pluripotent Stem Cells Cultured Under Feeder-Free Conditions

Muhammad Abid Sheikh¹, Farah Helena Afandi¹, Grazia Iannello², Barbara Corneo², Bright Starling Emerald^3,4,5, Suraiya Anjum Ansari^1,4,5

¹Department of Biochemistry and Molecular Biology, College of Medicine and Health Sciences, United Arab Emirates University, ²Columbia Stem Cell Initiative, Stem Cell Core, Columbia University Irving Medical Center, ³Department of Anatomy, College of Medicine and Health Sciences, United Arab Emirates University, ⁴Zayed Center for Health Sciences, United Arab Emirates University, ⁵ASPIRE Precision Medicine Research Institute Abu Dhabi (PMRI-AD), United Arab Emirates University

The presented method describes the generation of a CRISPR-mediated gene knockout in the human embryonic stem cell (hESC) line H9, which stably expresses sgRNAs targeting the L2HGDH gene using a highly efficient lentiviral-mediated gene delivery system.