Emory University School of Medicine 18 articles published in JoVE Biology Streamlined Intravital Imaging Approach for Long-Term Monitoring of Epithelial Tissue Dynamics on an Inverted Confocal Microscope Michael Hamersky IV1, Khushi Tekale1, L. Matthew Winfree2, Matthew J. M. Rowan1,3, Lindsey Seldin1,4,5,6 1Department of Cell Biology, Emory University School of Medicine, 2Independent scholar, 3Center for Neurodegenerative Disease, Emory University School of Medicine, 4Department of Dermatology, Emory University School of Medicine, 5Winship Cancer Institute, Emory University School of Medicine, 6Atlanta Veterans Affairs Medical Center The protocol presents a new tool to simplify intravital imaging using inverted confocal microscopy. Behavior Walk with Me Hybrid Virtual/In-Person Walking for Older Adults with Neurodegenerative Disease Meredith D. Wells1, Amy Morse2, Jolie Barter1, Kevin Mammino3, Allison A. Bay1, Todd Prusin1, Madeleine E. Hackney1,2,3,4,5 1Division of Geriatrics and Gerontology, Department of Medicine, Emory University School of Medicine, 2Department of Rehabilitation Medicine, Emory University School of Medicine, 3Atlanta VA Center for Visual and Neurocognitive Rehabilitation, 4Emory University School of Nursing, 5Birmingham/Atlanta VA Geriatric Research Education and Clinical Center The present protocol aims to inform rehabilitation specialists and fitness instructors in safe, feasible, and evidence-based methods of delivering virtual and in-person walking classes to older adults with neurodegenerative diseases. Neuroscience Systems Analysis of the Neuroinflammatory and Hemodynamic Response to Traumatic Brain Injury Rowan O. Brothers*1, Sara Bitarafan*2,3, Alyssa F. Pybus1,3, Levi B. Wood*1,2,3, Erin M. Buckley*1,4,5 1Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, 2George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology, 3Parker H. Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 4Department of Pediatrics, Emory University School of Medicine, 5 This protocol presents methods to characterize the neuroinflammatory and hemodynamic response to mild traumatic brain injury and to integrate these data as part of a multivariate systems analysis using partial least squares regression. Biochemistry Generation and Assembly of Virus-Specific Nucleocapsids of the Respiratory Syncytial Virus Yunrong Gao1, Claire Ogilvie1, Anirudh Raghavan1, Chloe Von Hoffmann1, Bo Liang1 1Department of Biochemistry, Emory University School of Medicine For in-depth mechanistic analysis of the respiratory syncytial virus (RSV) RNA synthesis, we report a protocol of utilizing the chaperone phosphoprotein (P) for coexpression of the RNA-free nucleoprotein (N0) for subsequent in vitro assembly of the virus-specific nucleocapsids (NCs). Medicine Segmentation and Linear Measurement for Body Composition Analysis using Slice-O-Matic and Horos Sean Steele1, Fangyi Lin1, Thien-Linh Le1, Alexandra Medline1, Michelle Higgins1, Alex Sandberg1, Sean Evans1, Gordon Hong2, Milton A. Williams3, Mehmet A. Bilen4, Sarah Psutka5, Kenneth Ogan1, Viraj A. Master1 1Department of Urology, Emory University School of Medicine, 2Northeast Ohio Medical University, 3Department of Urology, University of Alabama at Birmingham, 4Department of Medical Oncology, Winship Cancer Institute of Emory University, 5Department of Urology, University of Washington Segmentation and linear measurements quantify skeletal muscle mass and adipose tissues using Computed Tomography and/or Magnetic Resonance Imaging images. Here, we outline the use of Slice-O-Matic software and Horos image viewer for rapid and accurate analysis of body composition. These methods can provide important information for prognosis and risk stratification. Biology Single-Cell Multiplexed Fluorescence Imaging to Visualize Viral Nucleic Acids and Proteins and Monitor HIV, HTLV, HBV, HCV, Zika Virus, and Influenza Infection Raven Shah*1, Shuiyun Lan*1, Maritza N. Puray-Chavez2, Dandan Liu2, Philip R. Tedbury1, Stefan G. Sarafianos1 1Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, 2Department of Molecular Microbiology & Immunology, University of Missouri School of Medicine Presented here is a protocol for a fluorescence imaging approach, multiplex immunofluorescent cell-based detection of DNA, RNA, and protein (MICDDRP), a method capable of simultaneous fluorescence single-cell visualization of viral protein and nucleic acids of different type and strandedness. This approach can be applied to a diverse range of systems. Medicine An Image Guided Transapical Mitral Valve Leaflet Puncture Model of Controlled Volume Overload from Mitral Regurgitation in the Rat Daniella Corporan1, Takanori Kono1, Daisuke Onohara1, Muralidhar Padala1,2 1Structural Heart Research and Innovation Laboratory, Carlyle Fraser Heart Center, Emory University Hospital Midtown, 2Division of Cardiothoracic Surgery, Joseph P. Whitehead Department of Surgery, Emory University School of Medicine A rodent model of left heart volume overload from mitral regurgitation is reported. Mitral regurgitation of controlled severity is induced by advancing a needle of defined dimensions into the anterior leaflet of the mitral valve, in a beating heart, with ultrasound guidance. Biology A Uniform Shear Assay for Human Platelet and Cell Surface Receptors via Cone-plate Viscometry M. Edward Quach1,2, Anum K. Syed1,2, Renhao Li1,2 1 We describe an in-solution method to apply uniform shear to platelet surface receptors using cone-plate viscometry. This method may also be used more broadly to apply shear to other cell types and cell-fragments and need not target a specific ligand-receptor pair. Biochemistry Analyses of Mitochondrial Calcium Influx in Isolated Mitochondria and Cultured Cells Joshua T. Maxwell1, Chin-Hsien Tsai1, Tahmina A. Mohiuddin2, Jennifer Q. Kwong1 1Department of Pediatrics, Emory University School of Medicine, 2Emory College of Arts and Sciences, Emory University Here, we present two protocols for the measurement of mitochondrial Ca2+ influx in isolated mitochondria and cultured cells. For isolated mitochondria, we detail a plate reader-based Ca2+ import assay using the Ca2+ sensitive dye calcium green-5N. For cultured cells, we describe a confocal microscopy method using the Ca2+ dye Rhod-2/AM. Biochemistry Enrichment of Detergent-insoluble Protein Aggregates from Human Postmortem Brain Ian Diner1, Tram Nguyen1, Nicholas T. Seyfried1 1Department of Biochemistry, Emory School of Medicine An abbreviated fractionation protocol for the enrichment of detergent-insoluble protein aggregates from human postmortem brain is described. Engineering 3D Printing of Biomolecular Models for Research and Pedagogy Eduardo Da Veiga Beltrame1, James Tyrwhitt-Drake2, Ian Roy3, Raed Shalaby4, Jakob Suckale4, Daniel Pomeranz Krummel5 1Department of Physics, Brandeis University, 2Bioinformatics and Computational Biosciences Branch (BCBB), NIH/NIAID/OD/OSMO/OCICB, 3Library/LTS/MakerLab, Brandeis University, 4Interfaculty Institute of Biochemistry (IFIB), University of Tübingen, 5Winship Cancer Institute, Emory University School of Medicine Physical models of biomolecules can facilitate an understanding of their structure-function for the researcher, aid in communication between researchers, and serve as an educational tool in pedagogical endeavors. Here, we provide detailed guidance for the 3D printing of accurate models of biomolecules using fused filament fabrication desktop 3D printers. Biology Purification of Native Complexes for Structural Study Using a Tandem Affinity Tag Method Clarisse van der Feltz1, Daniel Pomeranz Krummel1,2 1Department of Biochemistry, Brandeis University, 2Winship Cancer Institute, Emory University School of Medicine The Tandem Affinity Purification (TAP) method has been used extensively to isolate native complexes from cellular extract, primarily eukaryotic, for proteomics. Here, we present a TAP method protocol optimized for purification of native complexes for structural studies. Immunology and Infection Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice Lauren E. Hudson1, Taryn P. Stewart1, Milo B. Fasken2, Anita H. Corbett2, Tracey J. Lamb1 1Department of Pediatrics, Emory University School of Medicine, 2Department of Biochemistry, Emory University School of Medicine Presented here is a unified description of techniques that can be used to develop, transform, administer, and test heterologous protein expression of the probiotic yeast Saccharomyces boulardii. Immunology and Infection Isolation of Exosomes from the Plasma of HIV-1 Positive Individuals Kateena Addae Konadu1, Ming Bo Huang1, William Roth1, Wendy Armstrong2, Michael Powell1, Francois Villinger3,4, Vincent Bond1 1Department of Microbiology, Biochemistry, Immunology, Morehouse School of Medicine, 2Department of Medicine, Emory University School of Medicine, 3Department of Pathology and Laboratory Medicine, Emory University School of Medicine, 4Yerkes National Primate Research Center Techniques describing a gradient procedure to separate exosomes from human immunodeficiency virus (HIV) particles are described. This procedure was used to isolate exosomes away from HIV particles in human plasma from HIV-infected individuals. The isolated exosomes were analyzed for cytokine/chemokine content. Medicine A Thrombotic Stroke Model Based On Transient Cerebral Hypoxia-ischemia Yu-Yo Sun1, Chia-Yi Kuan1 1Department of Pediatrics, Division of Neurology, The Emory University School of Medicine Thromboembolic stroke models are vital tools for optimizing the recanalization therapy. Here we report a murine thrombotic stroke model based on transient cerebral hypoxic-ischemic (tHI) insult, which triggers thrombosis and infarction, and responds favorably to tissue plasminogen activator (tPA)-mediated fibrinolysis in a therapeutic window similar to those in stroke patients. Behavior Community-based Adapted Tango Dancing for Individuals with Parkinson's Disease and Older Adults Madeleine E. Hackney1, Kathleen McKee2 1Center for Visual and Neurocognitive Rehabilitation, Atlanta VAMC, Department of Medicine, Division of General Medicine and Geriatrics, Emory University School of Medicine, 2 Adapted tango has demonstrated efficacy for improving mobility and balance. We describe the dissemination of adapted tango teaching methods to dance instructor trainees and the implementation of adapted tango by the trainees in the community for improving mobility and balance in older adults and individuals with Parkinson’s disease. Neuroscience An In Vitro Adult Mouse Muscle-nerve Preparation for Studying the Firing Properties of Muscle Afferents Joy A. Franco1, Heidi E. Kloefkorn2, Shawn Hochman3, Katherine A. Wilkinson4 1Department of Mechanical Engineering, San José State University, 2J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, 3Department of Physiology, Emory University School of Medicine, 4Department of Biological Sciences, San José State University Muscle sensory neurons are involved in proprioceptor signaling and also report on metabolic state and injury related events. We describe an adult mouse in vitro muscle-nerve preparation for studies on stretch-activated muscle afferents. Neuroscience Modeling Astrocytoma Pathogenesis In Vitro and In Vivo Using Cortical Astrocytes or Neural Stem Cells from Conditional, Genetically Engineered Mice Robert S. McNeill1, Ralf S. Schmid2, Ryan E. Bash3, Mark Vitucci4, Kristen K. White1, Andrea M. Werneke3, Brian H. Constance5, Byron Huff6, C. Ryan Miller2,3,7 1Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 2Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, 3Division of Neuropathology, Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 4Curriculum in Genetics and Molecular Biology, University of North Carolina School of Medicine, 5Biological and Biomedical Sciences Program, University of North Carolina School of Medicine, 6Department of Radiation Oncology, Emory University School of Medicine, 7Department of Neurology, Neurosciences Center, University of North Carolina School of Medicine Phenotypically wild-type astrocytes and neural stem cells harvested from mice engineered with floxed, conditional oncogenic alleles and transformed via viral Cre-mediated recombination can be used to model astrocytoma pathogenesis in vitro and in vivo by orthotopic injection of transformed cells into brains of syngeneic, immune-competent littermates.
