Ottawa Hospital Research Institute View Institution's Website 11 articles published in JoVE Immunology and Infection Detection of SARS-CoV-2 Receptor-Binding Domain Antibody using a HiBiT-Based Bioreporter Reza Rezaei1,2, Abera Surendran1,2, Ragunath Singaravelu1,2, Taylor R. Jamieson1,2, Parisa Taklifi3, Joanna Poutou1,2, Taha Azad1,2, Carolina S. Ilkow1,2 1Ottawa Hospital Research Institute, 2Department of Biochemistry, Microbiology and Immunology, University of Ottawa, 3Department of Biotechnology, University of Tehran The outlined protocol describes the procedure for producing the HiBiT-receptor-binding domain protein complex and its application for fast and sensitive detection of SARS-CoV-2 antibodies. Immunology and Infection Detection of SARS-CoV-2 Neutralizing Antibodies using High-Throughput Fluorescent Imaging of Pseudovirus Infection Taylor R. Jamieson1,2, Joanna Poutou1,2, Ricardo Marius1,2, Xiaohong He1, Reza Rezaei1,2, Taha Azad1,2, Carolina S. Ilkow1,2 1Ottawa Hospital Research Institute, 2Department of Biochemistry, Microbiology and Immunology, University of Ottawa The protocol described here outlines a fast and effective method for measuring neutralizing antibodies against the SARS-CoV-2 spike protein by evaluating the ability of convalescent serum samples to inhibit infection by an enhanced green fluorescent protein-labeled vesicular stomatitis virus pseudotyped with spike glycoprotein. Biology Isolation of CD146+ Resident Lung Mesenchymal Stromal Cells from Rat Lungs Jennifer J.P. Collins1,2, Marius A. Möbius1,3,4, Bernard Thébaud1,2,5 1Sinclair Centre for Regenerative Medicine, Ottawa Hospital Research Institute, 2University of Ottawa, 3Department of Neonatology and Pediatric Critical Care Medicine, Medical Faculty and University Hospital Carl Gustav Carus, Technische Universität Dresden, 4DFG Research Center and Cluster of Excellence for Regenerative Therapies (CRTD), Technische Universität, Dresden, 5 This protocol describes an isolation technique for obtaining primary lung resident mesenchymal stromal cells from rats, through the use of enzymatic digestion, density gradient separation, plastic adherence and CD146+ magnetic bead selection. Biology High-throughput Titration of Luciferase-expressing Recombinant Viruses Vanessa Garcia1,2, Ramya Krishnan1,2, Colin Davis1,2, Cory Batenchuk1,2, Fabrice Le Boeuf1,3, Hesham Abdelbary1,3, Jean-Simon Diallo1,2 1Center for Innovative Cancer Research, Ottawa Hospital Research Institute, 2Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, 3Faculty of Medicine, University of Ottawa This article presents a high-throughput luciferase expression-based method of titrating various RNA and DNA viruses using automated and manual liquid handlers. Medicine A Mouse Tumor Model of Surgical Stress to Explore the Mechanisms of Postoperative Immunosuppression and Evaluate Novel Perioperative Immunotherapies Lee-Hwa Tai1, Christiano Tanese de Souza1, Shalini Sahi1, Jiqing Zhang1,2,4, Almohanad A Alkayyal1,3,5, Abhirami Anu Ananth1,3, Rebecca A.C. Auer1,6 1Centre for Innovative Cancer Research, Ottawa Hospital Research Institute, 2Department of Cellular and Molecular Medicine, University of Ottawa, 3Department of Biochemistry, Microbiology and Immunology, University of Ottawa, 4Department of Neurosurgery, The Second Hospital of Shandong University, 5Department of Medical Laboratory Technology, University of Tabuk, 6Department of Surgery, Ottawa General Hospital A mouse tumor model of surgical stress is used to explore how postoperative immune suppression promotes metastatic disease and to evaluate immunostimulatory perioperative therapies. Neuroscience Dissection of the Transversus Abdominis Muscle for Whole-mount Neuromuscular Junction Analysis Lyndsay Murray1, Thomas H Gillingwater2, Rashmi Kothary1 1Regenerative Medicine Program, Ottawa Hospital Research Institute, 2Centre for Integrative Physiology, University of Edinburgh In this video we demonstrate a protocol for dissection of the transversus abdominis muscle of the mouse and use immunofluorescence and microscopy to visualize neuromuscular junctions. Biology Isolation and Culture of Individual Myofibers and their Satellite Cells from Adult Skeletal Muscle Alessandra Pasut1,2, Andrew E. Jones1,2, Michael A. Rudnicki1,2 1Sprott Center for Stem Cell Research, Ottawa Hospital Research Institute, 2Department of Cellular and Molecular Medicine, University of Ottawa Isolation and culture of myofibers is the gold standard in vitro system to study the transition of satellite cells through quiescence, activation and differentiation. Importantly, the single myofiber culture system preserves the myofiber/stem cell association, which is an essential component of the muscle stem cell niche. Neuroscience Derivation of Enriched Oligodendrocyte Cultures and Oligodendrocyte/Neuron Myelinating Co-cultures from Post-natal Murine Tissues Ryan W. O'Meara1,2, Scott D. Ryan1, Holly Colognato3, Rashmi Kothary1,2,4 1Regenerative Medicine Program, Ottawa Hospital Research Institute, 2Department of Cellular and Molecular Medicine, University of Ottawa, 3Department of Pharmacological Sciences, Stony Brook University, 4Department of Medicine, University of Ottawa This article describes methods to derive enriched populations of murine oligodendrocyte precursor cells (OPCs) in primary culture, which differentiate to produce mature oligodendrocytes (OLs). In addition, this report describes techniques to produce murine myelinating co-cultures by seeding mouse OPCs onto a neurite bed of mouse dorsal root ganglion neurons (DRGNs). Biology Lentiviral-mediated Knockdown During Ex Vivo Erythropoiesis of Human Hematopoietic Stem Cells Carmen G. Palii1, Roya Pasha1, Marjorie Brand1,2 1The Sprott Center for Stem Cell Research, Regenerative Medicine Program, Ottawa Hospital Research Institute, 2Department of Cellular and Molecular Medicine, University of Ottawa An ex vivo protocol to generate mature human red blood cells from hematopoietic stem/progenitors is described. Additionally we describe an efficient lentiviral-delivery method to knockdown the transcription factor TAL1 in primary erythroid cells. The efficiency of lentivirus mediated gene delivery is demonstrated using GFP expressing viruses. Medicine Ex Vivo Infection of Live Tissue with Oncolytic Viruses Jean-Simon Diallo1, Dominic Roy1, Hesham Abdelbary1, Naomi De Silva1, John C. Bell1 1Center for Innovative Cancer Research, Ottawa Hospital Research Institute (OHRI) Oncolytic viruses are promising for cancer therapeutics. The ability to ascertain the infectability of live tissue specimens obtained from patients prior to treatment is a unique advantage of this therapeutic approach. This protocol describes how to process tissues for ex vivo infection with oncolytic virus and subsequent viral quantification. Biology Immunohistochemistry on Paraffin Sections of Mouse Epidermis Using Fluorescent Antibodies Tammy-Claire Troy1, Azadeh Arabzadeh2, Adebola Enikanolaiye2, Nathalie Lariviere2, Kursad Turksen2 1Chronic Disease Program, Ottawa Health Research Institute, 2Department of Cellular and Molecular Medicine, Ottawa Health Research Institute We describe a reliable method for immunolocalization within the epidermis modified for both frozen and paraffin sections that we use very routinely in our laboratory.
