Children's Hospital of Pittsburgh of UPMC 5 articles published in JoVE Biology Studying the Epithelial Effects of Intestinal Inflammation In Vitro on Established Murine Colonoids Erin Crawford*1, Heather L. Mentrup*2,3, Elizabeth A. Novak2,3, Kevin P. Mollen2,3 1Division of Pediatric Surgery, UPMC Children’s Hospital of Pittsburgh, 3Department of Surgery, University of Pittsburgh School of Medicine We describe a protocol detailing the isolation of murine colonic crypts for the development of 3-dimensional colonoids. The established colonoids can then be terminally differentiated to reflect the cellular composition of the host epithelium prior to receiving an inflammatory challenge or being directed to establish an epithelial monolayer. Medicine Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery Joseph C. Fusco*1, Chen Congde*2, Xiangwei Xiao1, Krishna Prasadan1, David Ricks1, George K. Gittes1 1 Pancreatic duct infusion is an important technique that can allow for lineage tracing, gene introduction, and cell line-specific targeting. A pancreatic duct infusion technique for drug and viral delivery to pancreatic cells is presented here. Biology Guided Protocol for Fecal Microbial Characterization by 16S rRNA-Amplicon Sequencing Ayelet Di Segni1, Tzipi Braun1, Marina BenShoshan1, Sarit Farage Barhom1, Efrat Glick Saar1, Karen Cesarkas1, James E. Squires2, Nathan Keller1,3, Yael Haberman1,4 1Sheba Medical Center, 2 This manuscript describes a detailed standardized protocol of high-throughput 16S rRNA-amplicon sequencing. The protocol introduces an integrated, uniformed, feasible, and inexpensive protocol starting from fecal sample collection through data analyses. This protocol enables analysis of large numbers of samples with rigorous standards and several controls. Immunology and Infection Analysis of 18FDG PET/CT Imaging as a Tool for Studying Mycobacterium tuberculosis Infection and Treatment in Non-human Primates Alexander G. White*1, Pauline Maiello*1, M. Teresa Coleman1, Jaime A. Tomko1, L. James Frye1, Charles A. Scanga1, Philana Ling Lin2, JoAnne L. Flynn1 1Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, 2 Here, we present a protocol to describe the analysis of 18F-FDG PET/CT imaging in non-human primates that have been infected with M. tuberculosis to study disease process, drug treatment, and disease reactivation. Developmental Biology In Utero Intra-cardiac Tomato-lectin Injections on Mouse Embryos to Gauge Renal Blood Flow Christopher C. Rymer1,2, Sunder Sims-Lucas1,2 1 This manuscript describes a technique for visualization of the developing vasculature. Here we utilized in utero intra-cardiac FITC-labeled tomato lectin microinjections on mouse embryos. Using this technique, we delineate the perfused and unperfused vessels throughout the embryonic kidney.