Children's Hospital of Philadelphia

5 articles published in JoVE

Generation of High Quality Chromatin Immunoprecipitation DNA Template for High-throughput Sequencing (ChIP-seq)

JoVE 50286 4/19/2013

Sandra Deliard¹, Jianhua Zhao¹, Qianghua Xia¹, Struan F.A. Grant^1,2

¹Division of Human Genetics, Children's Hospital of Philadelphia Research Institute, ²Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania

The combination of chromatin immunoprecipitation and ultra-high-throughput sequencing (ChIP-seq) can identify and map protein-DNA interactions in a given tissue or cell line. Outlined is how to generate a high quality ChIP template for subsequent sequencing, using experience with the transcription factor TCF7L2 as an example.

Investigations on Alterations of Hippocampal Circuit Function Following Mild Traumatic Brain Injury

JoVE 4411 11/19/2012

Colin J. Smith^1,2, Brian N. Johnson¹, Jaclynn A. Elkind¹, Jill M. See¹, Guoxiang Xiong¹, Akiva S. Cohen^1,3

¹Division of Neurology, Children's Hospital of Philadelphia, ²Neuroscience Graduate Group, Perelman School of Medicine at the University of Pennsylvania, ³Department of Pediatrics, Perelman School of Medicine at the University of Pennsylvania

A multi-faceted approach to investigating functional changes to hippocampal circuitry is explained. Electrophysiological techniques are described along with the injury protocol, behavioral testing and regional dissection method. The combination of these techniques can be applied in similar fashion for other brain regions and scientific questions.

Generation of Human Induced Pluripotent Stem Cells from Peripheral Blood Using the STEMCCA Lentiviral Vector

JoVE 4327 10/31/2012

Andreia Gianotti Sommer¹, Sarah S. Rozelle¹, Spencer Sullivan², Jason A. Mills³, Seon-Mi Park¹, Brenden W. Smith¹, Amulya M. Iyer¹, Deborah L. French³, Darrell N. Kotton¹, Paul Gadue³, George J. Murphy¹, Gustavo Mostoslavsky¹

¹Center for Regenerative Medicine (CReM), Boston University School of Medicine, ²Department of Hematology, Children's Hospital of Philadelphia, ³Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia

Here we show a simple and effective protocol for the generation of human iPSCs from 3-4 ml of peripheral blood using a single lentiviral reprogramming vector. Reprogramming of readily available blood cells promises to accelerate the utilization of iPSC technology by making it accessible to a broader research community.

Isolation of Rat Portal Fibroblasts by In situ Liver Perfusion

JoVE 3669 6/29/2012

Jessica W. Wen¹, Abby L. Olsen², Maryna Perepelyuk², Rebecca G. Wells²

¹Division of Gastroenterology, Hepatology & Nutrition, Department of Pediatrics, The Children's Hospital of Philadelphia, ²Department of Medicine, University of Pennsylvania

A technique for isolating portal fibroblasts from rat liver is described. Livers are perfused and digested in situ with collagenase, followed by ex vivo digestion of the liver slurry and size selection of cells. This method provides a pure population of portal fibroblasts without the need for passage in culture.

Stable Isotopic Profiling of Intermediary Metabolic Flux in Developing and Adult Stage Caenorhabditis elegans

JoVE 2288 2/27/2011

Marni J. Falk^1,2, Meera Rao*¹, Julian Ostrovsky*¹, Evgueni Daikhin¹, Ilana Nissim¹, Marc Yudkoff^1,2

¹Department of Pediatrics, The Children's Hospital of Philadelphia, ²Department of Pediatrics, University of Pennsylvania

Stable isotopic profiling by gas chromatography mass spectrometric analysis of intermediary metabolic flux is described in the nematode, Caenorhabditis elegans. Methods are detailed for assessing isotopic enrichment in carbon dioxide, organic acids, and amino acids following isotope exposure either during development on agar plates or during adulthood in liquid culture.