21 articles published in JoVE

• Genetics

  
  Bidirectional Retroviral Integration Site PCR Methodology and Quantitative Data Analysis Workflow Gajendra W. Suryawanshi*^1,2, Song Xu*³, Yiming Xie¹, Tom Chou³, Namshin Kim⁴, Irvin S. Y. Chen^1,5, Sanggu Kim^{6 1}UCLA AIDS Institute, University of California at Los Angeles (UCLA), ²Department of Microbiology, Immunology, & Molecular Genetics, University of California at Los Angeles (UCLA), ³Departments of Biomathematics and Mathematics, University of California at Los Angeles (UCLA), ⁴Personalized Genomic Medicine Research Center, Division of Strategic Research Groups, Korea Research Institute of Bioscience and Biotechnology, ⁵Department of Medicine, University of California at Los Angeles (UCLA), ⁶Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University (OSU) This manuscript describes the experimental procedure and software analysis for a bidirectional integration site assay that can simultaneously analyze upstream and downstream vector-host junction DNA. Bidirectional PCR products can be used for any downstream sequencing platform. The resulting data are useful for a high-throughput, quantitative comparison of integrated DNA targets.

• Chemistry

  
  NMR Spectroscopy as a Robust Tool for the Rapid Evaluation of the Lipid Profile of Fish Oil Supplements Kathryn Williamson¹, Emmanuel Hatzakis^{1,2 1}Department of Food Science and Technology, The Ohio State University, ²Foods for Health Discovery Theme, The Ohio State University Here, high-resolution ¹H and ¹³C Nuclear Magnetic Resonance (NMR) spectroscopy was used as a rapid and reliable tool for quantitative and qualitative analysis of encapsulated fish oil supplements.

• Chemistry

  
  NMR Spectroscopy as a Robust Tool for the Rapid Evaluation of the Lipid Profile of Fish Oil Supplements Kathryn Williamson¹, Emmanuel Hatzakis^{1,2 1}Department of Food Science and Technology, The Ohio State University, ²Foods for Health Discovery Theme, The Ohio State University Here, high-resolution ¹H and ¹³C Nuclear Magnetic Resonance (NMR) spectroscopy was used as a rapid and reliable tool for quantitative and qualitative analysis of encapsulated fish oil supplements.

• Neuroscience

  
  Assaying DNA Damage in Hippocampal Neurons Using the Comet Assay Somaira Nowsheen¹, Fen Xia², Eddy S. Yang^{1,3 1}Department of Radiation Oncology, University of Alabama-Birmingham, ²Department of Radiation Oncology, The Ohio State University Medical School, ³Department of Cell Biology, and Pharmacology and Toxicology, Comprehensive Cancer Center, University of Alabama at Birmingham School of Medicine, University of Alabama-Birmingham The comet assay is an efficient way of detecting single- and double-strand breaks, including alkali-labile sites and DNA-DNA/DNA-protein cross-links on the DNA in all cells including hippocampal neurons. The method takes advantage of the differential migration of DNA in an electric field due to differences in amount of DNA damage.

• Biology

  
  Collection, Isolation and Enrichment of Naturally Occurring Magnetotactic Bacteria from the Environment Zachery Oestreicher¹, Steven K. Lower^1,2, Wei Lin³, Brian H. Lower^{2 1}School of Earth Sciences, The Ohio State University, ²School of Environment & Natural Resources, The Ohio State University, ³Institute of Geology and Geophysics, Chinese Academy of Sciences We demonstrate a method to collect magnetotactic bacteria (MTB) that can be applied to natural waters. MTB can be isolated and enriched from sediment samples using a relatively simple setup that takes advantage of the bacteria's natural magnetism. Isolated MTB can then be examined in detail using both light and electron microscopy.

• Biology

  
  Ex Vivo Assessment of Contractility, Fatigability and Alternans in Isolated Skeletal Muscles Ki Ho Park¹, Leticia Brotto², Oanh Lehoang¹, Marco Brotto², Jianjie Ma¹, Xiaoli Zhao^{1,3 1}Department of Physiology and Biophysics, UMDNJ-Robert Wood Johnson Medical School, ²Muscle Biology Research Group, University of Missouri-Kansas City, ³Pharmacology division, College of Pharmacy, DHLRI, Ohio State University We describe a method to directly measure muscle force, muscle power, contractile kinetics and fatigability of isolated skeletal muscles in an in vitro system using field stimulation. Valuable information on Ca²⁺ handling properties and contractile machinery of the muscle can be obtained using different stimulating protocols.

• Immunology and Infection

  
  Induction of Graft-versus-host Disease and In Vivo T Cell Monitoring Using an MHC-matched Murine Model Bryan A. Anthony¹, Gregg A. Hadley^{1 1}Department of Surgery, The Ohio State University Medical Center Murine bone marrow transplantation is a widely used technique to study immunological mechanisms governing graft-versus-host disease in humans. The ability to monitor T cell trafficking patterns in vivo allows for detailed analysis of the development and perpetuation of T cell responses during graft-versus-host disease.

• Biology

  
  Detection of Nitric Oxide and Superoxide Radical Anion by Electron Paramagnetic Resonance Spectroscopy from Cells using Spin Traps Bhavani Gopalakrishnan¹, Kevin M. Nash¹, Murugesan Velayutham¹, Frederick A. Villamena^{1,2 1}The Davis Heart and Lung Research Institute, The Ohio State University, ²Department of Pharmacology, College of Medicine, The Ohio State University Electron paramagnetic resonance (EPR) spectroscopy was employed to detect nitric oxide from bovine aortic endothelial cells and superoxide radical anion from human neutrophils using iron (II)-N-methyl-D-glucamine dithiocarbamate, Fe(MGD)2 and 5,5-dimethyl-1-pyroroline-N-oxide, DMPO, respectively.

• Medicine

  
  Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats Megan Ryan Detloff^1,2, Lesley C. Fisher¹, Rochelle J. Deibert¹, D. Michele Basso^{1 1}Center for Brain and Spinal Cord Repair, School of Allied Medical Professions, The Ohio State University, ²Spinal Cord Research Center, Drexel University College of Medicine We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

• Neuroscience

  
  Enabling High Grayscale Resolution Displays and Accurate Response Time Measurements on Conventional Computers Xiangrui Li¹, Zhong-Lin Lu^{2,3,4,5 1}Center for Cognitive and Behavioral Brain Imaging, The Ohio State University, ²Department of Psychology, University of Southern California, ³Biomedical Engineering, University of Southern California, ⁴Neuroscience Graduate Program, University of Southern California, ⁵Department of Psychology, The Ohio State University Conventional computer hardware can not generate visual stimuli with sufficiently high grayscale resolution and measure response times with sufficient accuracy. We describe how to use the VideoSwitcher to produce high-resolution monochromatic displays, and the RTbox to measure response times with high accuracy on conventional computer hardware.

• Neuroscience

  
  Cut-loading: A Useful Tool for Examining the Extent of Gap Junction Tracer Coupling Between Retinal Neurons Hee Joo Choi¹, Christophe P. Ribelayga², Stuart C. Mangel^{1 1}Department of Neuroscience, Ohio State University College of Medicine, ²Department of Ophthalmology and Visual Science, University of Texas Medical School An easy and convenient method to determine the extent of gap junction tracer coupling between retinal neurons is described. This technique enables one to investigate the function of the electrical synapses between neurons in the intact retina under different illumination conditions and at different times of the day and night.

• Medicine

  
  Method for Novel Anti-Cancer Drug Development using Tumor Explants of Surgical Specimens Kaushal Joshi¹, Habibe Demir¹, Ryosuke Yamada¹, Takeshi Miyazaki¹, Abhik Ray-Chaudhury², Ichiro Nakano^{1 1}Department of Neurological Surgery, The Ohio State University Medical Center, ²Department of Pathology, The Ohio State University Medical Center Here, we established a method for drug efficacy testing with surgical specimens of brain tumors, termed “tumor explant method”. With this method, we can evaluate drug efficacy without breaking the microenvironment of solid tumors. To validate reliability of this method, we describe representative data with our glioma specimen treated with the current first-line chemotherapeutic agent, temozolomide.

• Immunology and Infection

  
  Use of an Optical Trap for Study of Host-Pathogen Interactions for Dynamic Live Cell Imaging Jenny M. Tam¹, Carlos E. Castro², Robert J. W. Heath³, Michael K. Mansour¹, Michael L. Cardenas¹, Ramnik J. Xavier³, Matthew J. Lang⁴, Jatin M. Vyas^{1 1}Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Harvard Medical School, ²Department of Mechanical and Aerospace Engineering, The Ohio State University, ³Center for Computational and Integrative Biology, Massachusetts General Hospital, Harvard Medical School, ⁴Dept. of Chemical and Biomolecular Engineering, Vanderbilt University A method is described to individually select, manipulate, and image live pathogens using an optical trap coupled to a spinning disk microscope. The optical trap provides spatial and temporal control of organisms and places them adjacent to host cells. Fluorescence microscopy captures dynamic intercellular interactions with minimal perturbation to cells.

• Medicine

  
  A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation Erik J. Zmuda¹, Catherine A. Powell^1,2, Tsonwin Hai^{1,2,3 1}Molecular and Cellular Biochemistry, Center for Molecular Neurobiology, The Ohio State University, ²Integrated Biomedical Science Graduate Program, The Ohio State University, ³Comprehensive Cancer Center, The Ohio State University Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

• Biology

  
  Identifying the Effects of BRCA1 Mutations on Homologous Recombination using Cells that Express Endogenous Wild-type BRCA1 Jeffrey Parvin¹, Natsuko Chiba², Derek Ransburgh^{1 1}Department of Biomedical Informatics, The Ohio State University, ²Departments of Molecular Immunology and Clinical Oncology, Tohoku University We provide a method for testing BRCA1 variants in a tissue culture based assay for homologous recombination repair of DNA damage by depleting endogenous BRCA1 protein from a cell using RNAi and replacing it with a BRCA1 point mutant that contains a coding change.

• Medicine

  
  Dual-mode Imaging of Cutaneous Tissue Oxygenation and Vascular Function Ronald X. Xu¹, Kun Huang², Ruogu Qin¹, Jiwei Huang¹, Jeff S. Xu¹, Liya Ding², Urmila S. Gnyawali³, Gayle M. Gordillo³, Surya C. Gnyawali^3,4, Chandan K. Sen^{3 1}Department of Biomedical Engineering, The Ohio State University, ²Department of Biomedical Informatics, The Ohio State University, ³Comprehensive Wound Center, The Ohio State University, ⁴Department of Surgery, The Ohio State University A dual-mode imaging system was developed for non-contact assessment of cutaneous tissue oxygenation and vascular function.

• Medicine

  
  High-frequency High-resolution Echocardiography: First Evidence on Non-invasive Repeated Measure of Myocardial Strain, Contractility, and Mitral Regurgitation in the Ischemia-reperfused Murine Heart Surya C. Gnyawali^1,2, Sashwati Roy^1,2, Jason Driggs^1,2, Savita Khanna^1,2, Thomas Ryan^2,3, Chandan K. Sen^{2 1}Department of Surgery, The Ohio State University, ²Heart and Lung Research Institute, The Ohio State University, ³Department of Cardiovascular Medicine, The Ohio State University High frequency Doppler ultrasound is a novel technology for assessing regional myocardial function. This work presents first evidence demonstrating applicability of this versatile imaging platform for the repeated measure of myocardial strain, dp/dt, and mitral regurgitation in the ischemia-reperfused (IR) murine heart.

• Biology

  
  Robotics and Dynamic Image Analysis for Studies of Gene Expression in Plant Tissues Carlos M. Hernandez-Garcia¹, Joseph M. Chiera^1,2, John J. Finer^{1 1}Department of Horticulture and Crop Science, The Ohio State University, ²Department of Plant Pathology, North Carolina State University We report a method for introduction, tracking and quantitative analysis of GFP expression in plant cells. This method utilizes a custom-designed robotics system for semi-continuous image collection from large numbers of samples, over time. We also demonstrate the use of ImageJ and ImageReady for analysis of image series.

• Biology

  
  Analysis of Gene Expression in Emerald Ash Borer (Agrilus planipennis) Using Quantitative Real Time-PCR Binny Bhandary¹, Swapna Priya Rajarapu¹, Loren Rivera-Vega¹, Omprakash Mittapalli^{1 1}Department of Entomology, The Ohio State University Quantitative real-time PCR (qRT-PCR) is an effective tool to diagnose mRNA levels in different insect tissues and developmental stages. In this report we show the use of qRT-PCR to ascertain mRNA levels in different larval tissues and developmental stages of the invasive insect species, emerald ash borer.

• Biology

  
  Tomato Analyzer: A Useful Software Application to Collect Accurate and Detailed Morphological and Colorimetric Data from Two-dimensional Objects Gustavo R. Rodríguez¹, Jennifer B. Moyseenko¹, Matthew D. Robbins¹, Nancy Huarachi Morejón¹, David M. Francis¹, Esther van der Knaap^{1 1}Department of Horticulture and Crop Science, The Ohio State University Tomato Analyzer (TA) quantifies attributes of two dimensional shapes and color in a reproducible and accurate manner. A step-by-step procedure for obtaining high quality digitalized images of tomato fruit, morphological and color analyses of these images and several applications using the data generated through this software are described.

• Biology

  
  Murine Renal Transplantation Procedure Jiao-Jing Wang¹, Sara Hockenheimer², Alice A. Bickerstaff¹, Gregg A. Hadley^{1 1}Department of Surgery, The Ohio State University, ²Integrated Biomedical Sciences Graduate Program, The Ohio State University Renal transplantation in mice is a technically challenging procedure that requires careful post-operative care and treatment for success.