St. Jude Children's Research Hospital

11 articles published in JoVE

• Immunology and Infection

  
  Evaluation of Caspase Activation to Assess Innate Immune Cell Death Joo-Hui Han¹, Rebecca E. Tweedell¹, Thirumala-Devi Kanneganti^{1 1} This protocol describes a comprehensive method for assessing caspase activation (caspase-1, caspase-3, caspase-7, caspase-8, caspase-9, and caspase-11) in response to both in vitro and in vivo (in mice) models of infection, sterile insults, and cancer to determine the initiation of cell death pathways, such as pyroptosis, apoptosis, necroptosis, and PANoptosis.

• Biochemistry

  
  JUMPn: A Streamlined Application for Protein Co-Expression Clustering and Network Analysis in Proteomics David Vanderwall¹, Poudel Suresh^1,2, Yingxue Fu², Ji-Hoon Cho², Timothy I. Shaw^2,3, Ashutosh Mishra², Anthony A. High², Junmin Peng^1,2, Yuxin Li^{1,2 1}Center for Proteomics and Metabolomics, St. Jude Children’s Research Hospital, ³ We present a systems biology tool JUMPn to perform and visualize network analysis for quantitative proteomics data, with a detailed protocol including data pre-processing, co-expression clustering, pathway enrichment, and protein-protein interaction network analysis.

• Biochemistry

  
  Quantification of Coenzyme A in Cells and Tissues Matthew W. Frank¹, Chitra Subramanian¹, Charles O. Rock¹, Suzanne Jackowski^{1 1} This method describes sample preparation from cultured cells and animal tissues, extraction and derivatization of coenzyme A in the samples, followed by high pressure liquid chromatography for purification and quantification of the derivatized coenzyme A by absorbance or fluorescence detection.

• Biology

  
  Characterization of MLKL-mediated Plasma Membrane Rupture in Necroptosis Dan E. McNamara*^1,2, Giovanni Quarato*³, Cliff S. Guy³, Douglas R. Green³, Tudor Moldoveanu^{1,2 1} We report methods for characterization of MLKL-mediated plasma membrane rupture in necroptosis including conventional and confocal live-cell microscopy imaging, scanning electron microscopy, and NMR-based lipid binding.

• Biochemistry

  
  Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification Anthony A. High¹, Haiyan Tan¹, Vishwajeeth R. Pagala¹, Mingming Niu^2,3, Ji-Hoon Cho¹, Xusheng Wang¹, Bing Bai², Junmin Peng^{1,2 1} We present a protocol to accurately quantitate proteins with isobaric labelling, extensive fractionation, bioinformatics tools, and quality control steps in combination with liquid chromatography interfaced to a high-resolution mass spectrometer.

• Medicine

  
  A Cell Culture Model of Resistance Arteries Lauren A. Biwer^1,2, Christophe Lechauve³, Sheri Vanhoose⁴, Mitchell J. Weiss³, Brant E. Isakson^{1,2 1}Department of Molecular Physiology and Biophysics, University of Virginia School of Medicine, ²Robert M. Berne Cardiovascular Research Center, University of Virginia School of Medicine, ³ A cell culture model of resistance arteries is described, allowing for the dissection of signaling pathways in endothelium, smooth muscle, or between endothelium and smooth muscle (the myoendothelial junction). The selective application of agonists or protein isolation, electron microscopy, or immunofluorescence can be utilized using this cell culture model.

• Medicine

  
  Rose Bengal Photothrombosis by Confocal Optical Imaging In Vivo: A Model of Single Vessel Stroke Lora Talley Watts¹, Wei Zheng¹, R. Justin Garling², Victoria C. Frohlich³, James Donald Lechleiter^{1 1}Department of Cellular and Structural Biology, The University of Texas Health Science Center San Antonio, ²School of Medicine, The University of Texas Health Science Center San Antonio, ³ Here, we describe a semi-invasive optical microscopy approach for the induction of a Rose Bengal photothrombotic clot in the somatosensory cortex of a mouse in vivo. The technical aspects of the imaging procedure are described from induction of a photothrombotic event to application and data collection.

• Neuroscience

  
  Mitochondria-associated ER Membranes (MAMs) and Glycosphingolipid Enriched Microdomains (GEMs): Isolation from Mouse Brain Ida Annunziata*¹, Annette Patterson*¹, Alessandra d'Azzo^{1 1}Department of Genetics, St Jude Children's Research Hospital This procedure illustrates how to isolate from the adult mouse brain the mitochondria-associated ER membranes or MAMs and the glycosphingolipid-enriched microdomain fractions from MAMs and mitochondrial preparations.

• Neuroscience

  
  Dissection and Imaging of Active Zones in the Drosophila Neuromuscular Junction Rebecca Smith¹, J. Paul Taylor^{1 1}Developmental Neurobiology, St. Jude Children’s Research Hospital The neuromuscular junction (NMJ) of Drosophila melanogaster is an important model system for studying normal synaptic function as well as perturbations to synaptic function found in certain neurological diseases. We present a protocol for dissection of the Drosophila larval motor system and immunostaining for active zone proteins within the NMJ.

• Medicine

  
  Guidelines for Elective Pediatric Fiberoptic Intubation Roland N. Kaddoum¹, Zulfiqar Ahmed², Alan A. D'Augsutine², Maria M. Zestos^{3 1}Department of Anesthesia, St. Jude Children's Research Hospital, ²Department of Anesthesia, Children's Hospital of Michigan, ³Department of Anesthesiology, Children's Hospital of Michigan We describe guidelines to perform a safe and efficient elective fiberoptic intubation in pediatric patients while maintaining spontaneous ventilation.

• Medicine

  
  A Novel Rescue Technique for Difficult Intubation and Difficult Ventilation Maria M. Zestos¹, Dima Daaboul¹, Zulfiqar Ahmed¹, Nasser Durgham¹, Roland Kaddoum^{2 1}Department of Anesthesia, Children’s Hospital of Michigan, ²Department of Anesthesia, St. Jude Children’s Research Hospital We describe a technique to maintain oxygenation and ventilation using an endotracheal tube inserted nasally to the level of the naso-pharynx while sealing the mouth and nares for successful positive pressure ventilation.