Ohio State University View Institution's Website 80 articles published in JoVE Bioengineering Automated Compression Testing of the Ocular Lens Debae Alzoubi1, Wade Rich1, Matthew A. Reilly1,2 1Department of Biomedical Engineering, The Ohio State University, 2Department of Ophthalmology & Visual Sciences, The Ohio State University We present an automated method for characterizing the effective elastic modulus of an ocular lens using a compression test. Medicine Successful Orthotopic Liver Transplantation in Mice Utilizing Microcomputed Tomography Angiography Qiang Zeng1, Doug A. Gouchoe2,3, Mahboubeh Nabavinia4, Yong Gyu Lee2, Xi Wang2, Terri A. Shaffer5, Mitchel R. Stacy4,5,6, Blake R. Peterson7,8, Bryan A. Whitson2,9, Christopher Breuer1,4, Sylvester M. Black1,10 1Center for Regenerative Medicine, The Research Institute at Nationwide Children’s Hospital, 5Division of Vascular Diseases and Surgery, The Ohio State University College of Medicine, 6Interdisciplinary Biophysics Graduate Program, The Ohio State University, 7Division of Medicinal Chemistry and Pharmacognosy, The Ohio State University College of Pharmacy, 8The Ohio State University Comprehensive Cancer Center, 9Division of Cardiac Surgery, Department of Surgery, The Ohio State Wexner Medical Center, 10Division of Transplantation, Department of Surgery, The Ohio State Wexner Medical Center In this protocol, we discuss the implementation of a model of successful orthotopic liver transplantation (OLT) in mice. Additionally, adjuvants to further analyze allograft patency after successful OLT in a mouse are discussed as well, specifically utilizing microcomputed tomography (microCT) scans. Medicine A Simple and Effective Method to Consistently Isolate Mouse Cardiomyocytes Sarah L. Sturgill1, Lorien G. Salyer1, Brandon J. Biesiadecki1, Mark T. Ziolo1 1Department of Physiology and Cell Biology, The Ohio State University The gold standard in cardiology for cellular and molecular functional experiments are cardiomyocytes. This article describes adaptations to the non-Langendorff technique to isolate mouse cardiomyocytes. Immunology and Infection Standardized In vitro Assays to Visualize and Quantify Interactions between Human Neutrophils and Staphylococcus aureus Biofilms Pranav S. J. B. Rana1, Erin S. Gloag2, Daniel J. Wozniak1,2 1Department of Microbiology, The Ohio State University, 2Department of Microbial Infection and Immunity, The Ohio State University The present protocol describes the study of neutrophil-biofilm interactions. Staphylococcus aureus biofilms are established in vitro and incubated with peripheral blood-derived human neutrophils. The oxidative burst response from neutrophils is quantified, and the neutrophil localization within the biofilm is determined by microscopy. Biochemistry Uracil-DNA Glycosylase Assay by Matrix-assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry Analysis Hui-Lan Chang*1, Kang-Yi Su*1,2, Steven D. Goodman3, Wern-Cherng Cheng2, Liang-In Lin1,2, Ya-Chien Yang1,2, Sui-Yuan Chang1,2, Woei-horng Fang1,2 1Department of Clinical Laboratory Sciences and Medical Biotechnology, College of Medicine, National Taiwan University, 2Department of Laboratory Medicine, National Taiwan University Hospital, 3Center for Microbial Pathogenesis, Nationwide Children’s Hospital and the Department of Pediatrics, The Ohio State University A non-labeled, non-radio-isotopic method to assay uracil-DNA glycosylase activity was developed using MALDI-TOF mass spectrometry for direct apurinic/apyrimidinic site-containing product analysis. The assay proved to be quite simple, specific, rapid, and easy to use for DNA glycosylase measurement. Biochemistry Extracellular Glucose Depletion as an Indirect Measure of Glucose Uptake in Cells and Tissues Ex Vivo Shashi Bhushan Kumar1, Shanvanth Arnipalli1, Adham Abushukur1, Silvia Carrau1, Priyanka Mehta1, Ouliana Ziouzenkova1 1Department of Human Sciences, The Ohio State University Extracellular depletion of fluorescently labeled glucose correlates with glucose uptake and could be used for high-throughput screening of glucose uptake in excised organs and cell cultures. Medicine Biological Preparation and Mechanical Technique for Determining Viscoelastic Properties of Zonular Fibers Juan Rodriguez1,2, Matthew Reilly3, Robert P. Mecham4, Steven Bassnett2,4 1Department of Basic Sciences, University of Health Sciences and Pharmacy, 2Department of Ophthalmology & Visual Sciences, Washington University School of Medicine, 3Department of Biomedical Engineering, The Ohio State University, 4Department of Cell Biology & Physiology, Washington University School of Medicine The protocol describes a method for the study of extracellular matrix viscoelasticity and its dependence on protein composition or environmental factors. The matrix system targeted is the mouse zonule. The performance of the method is demonstrated by comparing the viscoelastic behavior of wild-type zonular fibers with those lacking microfibril-associated glycoprotein-1. Medicine A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes Yong Gyu Lee1,2, Jung-Lye Kim1,2, Andre F. Palmer3, Brenda F. Reader4, Jianjie Ma1, Sylvester M. Black2,4, Bryan A. Whitson1,2 1Department of Surgery Division of Cardiac Surgery, The Ohio State University Wexner Medical Center, 2The Collaboration for Organ Perfusion, Protection, Engineering and Regeneration (COPPER) Laboratory, The Ohio State University College of Medicine, 3William G. Lowrie Department of Chemical and Biomolecular Engineering, College of Engineering, The Ohio State University, 4Comprehensive Transplant Center, The Ohio State University Wexner Medical Center Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection. Genetics Use of the Pyrimidine Analog, 5-Iodo-2′-Deoxyuridine (IdU) with Cell Cycle Markers to Establish Cell Cycle Phases in a Mass Cytometry Platform Raymond D Devine1, Gregory K Behbehani1 1Ohio State Comprehensive Cancer Center, Division of Hematology, The Ohio State University This protocol adapts cell cycle measurements for use in a mass cytometry platform. With the multi-parameter capabilities of mass cytometry, direct measurement of iodine incorporation allows identification of cells in S-phase while intracellular cycling markers enable characterization of each cell cycle state in a range of experimental conditions. Engineering Surgery and Sample Processing for Correlative Imaging of the Murine Pulmonary Valve Yifei Liu1, Yong-Ung Lee2, Tai Yi2, Ken Wu3, Cedric Bouchet-Marquis3, Han Chan3, Christopher K. Breuer2, David W. McComb1 1Center for Electron Microscopy and Analysis, The Department of Materials Science and Engineering, Ohio State University, 2 Here, we describe a correlative workflow for the excision, pressurization, fixation, and imaging of the murine pulmonary valve to determine the gross conformation and local extracellular matrix structures. Genetics Direct Reprogramming of Human Fibroblasts into Myoblasts to Investigate Therapies for Neuromuscular Disorders Camila F. Almeida1, Emma C. Frair1, Nianyuan Huang1, Reid Neinast2, Kim L. McBride2,3,4,6, Robert B. Weiss5, Kevin M. Flanigan1,6, Nicolas Wein1,6 1Center for Cardiovascular Research, The Research Institute at Nationwide Children’s Hospital, 3Division of Genetic and Genomic Medicine, Nationwide Children’s Hospital, 5Department of Human Genetics, The University of Utah School of Medicine, 6Department of Pediatrics, The Ohio State University This protocol describes the conversion of skin fibroblasts into myoblasts and their differentiation into myotubes. The cell lines are derived from patients with neuromuscular disorders and can be used to investigate pathological mechanisms and to test therapeutic strategies. Developmental Biology Generation and Expansion of Human Cardiomyocytes from Patient Peripheral Blood Mononuclear Cells Shiqiao Ye1,2, Xiaoping Wan3, Juan Su1,2, Akshar Patel1,2,4, Blake Justis1,2,5, Isabelle Deschênes3, Ming-Tao Zhao1,2,5,6 1The Heart Center, Nationwide Children’s Hospital, 3Department of Physiology and Cell Biology, The Ohio State University College of Medicine, 4Department of Anatomy, The Ohio State University College of Medicine, 5MCDB Graduate Program, The Ohio State University, 6Department of Pediatrics, The Ohio State University College of Medicine Here, we present a protocol to robustly generate and expand human cardiomyocytes from patient peripheral blood mononuclear cells. Biology Preparing and Injecting Embryos of Culex Mosquitoes to Generate Null Mutations using CRISPR/Cas9 Megan E. Meuti1, Robert Harrell2 1Department of Entomology, The Ohio State University, 2Insect Transformation Facility, Institute for Bioscience and Biotechnology Research, University of Maryland CRISPR/Cas9 is increasingly used to characterize gene function in non-model organisms. This protocol describes how to generate knock-out lines of Culex pipiens, from preparing injection mixes, to obtaining and injecting mosquito embryos, as well as how to rear, cross, and screen injected mosquitoes and their progeny for desired mutations. Neuroscience Imaging and Analysis of Neurofilament Transport in Excised Mouse Tibial Nerve Nicholas P. Boyer1, Maite Azcorra1,2, Peter Jung3,4, Anthony Brown1 1Department of Neuroscience, The Ohio State University, 2Present address: Interdepartmental Neuroscience Graduate Program and Department of Neurobiology, Northwestern University, 3Quantitative Biology Institute, Ohio University, 4Department of Physics and Astronomy, Ohio University We describe fluorescence photoactivation methods to analyze the axonal transport of neurofilaments in single myelinated axons of peripheral nerves from transgenic mice that express a photoactivatable neurofilament protein. Immunology and Infection Propagation of the Microsporidian Parasite Edhazardia aedis in Aedes aegypti Mosquitoes Anthony Grigsby*1, Brendan J. Kelly*1, Neil D. Sanscrainte2, James J. Becnel2, Sarah M. Short1 1Department of Entomology, The Ohio State University, 2USDA/ARS Center for Medical, Agricultural, and Veterinary Entomology A protocol to culture the microsporidian parasite Edhazardia aedis. The parasite is passaged from one generation of Aedes aegypti mosquitoes to the next via horizontal transfer at the larval stage followed by vertical transmission at the adult stage. Live sporoplasms survive long-term in infected eggs. Biology A 3D Printed Pollen Trap for Bumble Bee (Bombus) Hive Entrances Houston J. Judd1,2, Craig Huntzinger2, Ricardo Ramirez1, James P. Strange2,3 1Department of Biology, Utah State University, 2USDA-ARS Pollinating Insect Research Unit, 3Department of Entomology, The Ohio State University We present a non-lethal and automated mechanism to collect pollen from bumble bee (Bombus) workers returning to a hive. Instructions for producing, preparing, installing and using the devices are included. By using 3D-printed objects, modification to the design was timely, efficient and allowed for quick turnaround for testing. Cancer Research Mapping the Structure-Function Relationships of Disordered Oncogenic Transcription Factors Using Transcriptomic Analysis Iftekhar A. Showpnil1,2, Kyle R. Miller1, Cenny Taslim1, Kathleen I. Pishas1, Stephen L. Lessnick1,3, Emily R. Theisen1,4 1 Intrinsically disordered domains are important for oncogenic fusion transcription factor function. To therapeutically target these proteins, a more detailed understanding of the regulatory mechanisms employed by these domains is required. Here, we use transcriptomics to map important structural features of the intrinsically disordered EWS domain in Ewing sarcoma. Cancer Research Modeling Brain Metastases Through Intracranial Injection and Magnetic Resonance Imaging Jennifer A. Geisler1,2,3, Jonathan M. Spehar1,2, Sarah A. Steck1,2, Anna Bratasz1,4, Reena Shakya1, Kimerly Powell1,4, Gina M. Sizemore1,2 1The Comprehensive Cancer Center, The Ohio State University, 2Department of Radiation Oncology, The Ohio State University, 3Department of Veterinary Biosciences, The Ohio State University, 4Davis Heart & Lung Research Institute, The Ohio State University Intracranial brain metastasis modeling is complicated by an inability to monitor tumor size and response to treatment with precise and timely methods. The presented methodology couples intracranial tumor injection with magnetic resonance imaging analysis, which when combined, cultivates precise and consistent injections, enhanced animal monitoring, and accurate tumor volume measurements. Cancer Research Pathological Analysis of Lung Metastasis Following Lateral Tail-Vein Injection of Tumor Cells Katie A Thies1, Sarah Steck1, Sue E Knoblaugh2, Steven T Sizemore1 1Department of Radiation Oncology, Arthur G. James Comprehensive Cancer Center and Richard L. Solove Research Institute, The Ohio State University Medical Center, 2Department of Veterinary Biosciences, Comparative Pathology and Digital Imaging Shared Resource, The Ohio State University Intravenous injection of cancer cells is often used in metastasis research, but the metastatic tumor burden can be difficult to analyze. Herein, we demonstrate a tail-vein injection model of metastasis and include a novel approach to analyze the resulting metastatic lung tumor burden. Medicine Repeated Orotracheal Intubation in Mice Andrew M. Nelson1, Katherine E. Nolan1, Ian C. Davis1 1Department of Veterinary Biosciences, The Ohio State University The goal of this article is to describe a refined method of intubation of the laboratory mouse. The method is noninvasive and, therefore, ideal for studies that require serial monitoring of respiratory function and/or instillation of treatments into the lung. Bioengineering Bioparticle Microarrays for Chemotactic and Molecular Analysis of Human Neutrophil Swarming in vitro Nicole Walters1, Eduardo Reátegui1,2 1William G. Lowrie Department of Chemical and Biomolecular Engineering, The Ohio State University, 2Comprehensive Cancer Center, The Ohio State University This protocol generates bioparticle microarrays that provide spatially controlled neutrophil swarming. It provides easy access to the mediators that neutrophils release during migration and allows for quantitative imaging analysis. Chemistry Quantification of Metal Leaching in Immobilized Metal Affinity Chromatography Coleman M. Swaim1, Tyler J. Brittain1, Daniel R. Marzolf2, Oleksandr Kokhan1 1Department of Chemistry and Biochemistry, James Madison University, 2Biophysics Graduate Program, Ohio State University We present an assay for easy quantification of metals introduced to samples prepared using immobilized metal affinity chromatography. The method uses hydroxynaphthol blue as the colorimetric metal indicator and a UV-Vis spectrophotometer as the detector. Engineering Multimodal 3D Printing of Phantoms to Simulate Biological Tissue Canzhen Ma*1,2, Shuwei Shen*1,2, Guangli Liu1, Siyue Guo1,2, Buyun Guo1,2, Jialuo Li1,2, Kuiming Huang1,2, Yidan Zheng1, Pengfei Shao1,2, Erbao Dong1,2, Jiaru Chu1, Ronald X. Xu1,2,3 1Department of Precision Machinery and Precision Instrumentation, University of Science and Technology of China, 2Key Laboratory of Precision Scientific Instrumentation of Anhui Higher Education Institutes, University of Science and Technology of China, 3Department of Biomedical Engineering, The Ohio State University Spin coating, polyjet printing, and fused deposition modeling are integrated to produce multilayered heterogeneous phantoms that simulate structural and functional properties of biological tissue. Biochemistry Expression and Purification of Nuclease-Free Oxygen Scavenger Protocatechuate 3,4-Dioxygenase Ryan K. Messer1, Miguel A. Lopez Jr.1, Gayan Senavirathne1, Kristine E. Yoder1 1Cancer Biology and Genetics, College of Medicine, The Ohio State University Protocatechuate 3,4-dioxygenase (PCD) can enzymatically remove free diatomic oxygen from an aqueous system using its substrate protocatechuic acid (PCA). This protocol describes the expression, purification, and activity analysis of this oxygen scavenging enzyme. Genetics Identification of Footprints of RNA:Protein Complexes via RNA Immunoprecipitation in Tandem Followed by Sequencing (RIPiT-Seq) Lauren Woodward1, Pooja Gangras1, Guramrit Singh1 1Department of Molecular Genetics, Center for RNA Biology, The Ohio State University Here, we present a protocol to enrich endogenous RNA binding sites or "footprints" of RNA:protein (RNP) complexes from mammalian cells. This approach involves two immunoprecipitations of RNP subunits and is therefore dubbed RNA immunoprecipitation in tandem (RIPiT). Cancer Research Rapid Generation of Primary Murine Melanocyte and Fibroblast Cultures Brandon M. Murphy1, Tirzah J. Weiss1, Christin E. Burd1,2 1Department of Cancer Biology and Genetics, The Ohio State University, 2Department of Molecular Genetics, The Ohio State University This protocol outlines a rapid method to simultaneously generate melanocyte and fibroblast cultures from the skin of 0-4 day old mice. These primary cultures can be maintained and manipulated in vitro to study a variety of physiologically relevant processes, including skin cell biology, pigmentation, wound healing and melanoma. Immunology and Infection Evaluation of Host-Pathogen Responses and Vaccine Efficacy in Mice Kyle Caution1, Kacy Yount1, Rajendar Deora1,2, Purnima Dubey1 1Department of Microbial Infection & Immunity, The Ohio State University, 2Department of Microbiology, The Ohio State University Here we present an elegant protocol for in vivo evaluation of vaccine effectiveness and host immune responses. This protocol can be adapted for vaccine models that study viral, bacterial, or parasitic pathogens. Bioengineering Measuring Global Cellular Matrix Metalloproteinase and Metabolic Activity in 3D Hydrogels Abdulaziz S. Fakhouri1,2,3, Jennifer L. Leight1,2 1Department of Biomedical Engineering, The Ohio State University, 2The Ohio State University Comprehensive Cancer Center, Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, 3Biomedical Technology Department, King Saud University Here, a protocol is presented for encapsulating and culturing cells in poly(ethylene glycol) (PEG) hydrogels functionalized with a fluorogenic matrix metalloproteinase (MMP)-degradable peptide. Cellular MMP and metabolic activity are measured directly from the hydrogel cultures using a standard microplate reader. Cancer Research Detection of Protease Activity by Fluorescent Peptide Zymography Ameya A. Deshmukh1,2, Jessica L. Weist1, Jennifer L. Leight1,2 1Comprehensive Cancer Center, James Cancer Hospital and Solove Research Center, Ohio State University, 2Department of Biomedical Engineering, College of Engineering, Ohio State University Here, we present a detailed protocol for a modified zymographic technique in which fluorescent peptides are used as the degradable substrate in place of native proteins. Electrophoresis of biological samples in fluorescent peptide zymograms enables detection of a wider range of proteases than previous zymographic techniques. Immunology and Infection High-throughput Measurement of Plasma Membrane Resealing Efficiency in Mammalian Cells Jonathan G.T. Lam1,2,3, Chi Song4, Stephanie Seveau1,2,3 1Department of Microbial Infection and Immunity, The Ohio State University, 2Department of Microbiology, The Ohio State University, 3Infectious Diseases Institute, The Ohio State University, 4Division of Biostatistics, College of Public Health, The Ohio State University Here we describe a high-throughput fluorescence-based assay that measures the plasma membrane resealing efficiency through fluorometric and imaging analyses in living cells. This assay can be used for screening drugs or target genes that regulate plasma membrane resealing in mammalian cells. Behavior Novel Object Recognition and Object Location Behavioral Testing in Mice on a Budget Jiyeon K. Denninger1, Bryon M. Smith1, Elizabeth D. Kirby1,2,3 1Department of Psychology, Ohio State University, 2Department of Neuroscience, Ohio State University, 3Center for Chronic Brain Injury, Ohio State University Here we provide a protocol which includes comprehensive instructions for the economical establishment of murine object location and novel object recognition behavioral testing, including the design, cost, and construction of required equipment as well as execution of behavioral testing, data collection, and analysis. Immunology and Infection The Galleria mellonella Waxworm Infection Model for Disseminated Candidiasis Matthew J. Dunn1, Andrew L. Woodruff1, Matthew Z. Anderson1,2 1Department of Microbiology, The Ohio State University, 2Department of Microbial Infection and Immunity, The Ohio State University Galleria mellonella serves as an invertebrate model for disseminated candidiasis. Here, we detail the infection protocol and provide supporting data for the model's effectiveness. Behavior Gaze in Action: Head-mounted Eye Tracking of Children's Dynamic Visual Attention During Naturalistic Behavior Lauren K. Slone1, Drew H. Abney1, Jeremy I. Borjon1, Chi-hsin Chen2, John M. Franchak3, Daniel Pearcy1, Catalina Suarez-Rivera1, Tian Linger Xu1, Yayun Zhang1, Linda B. Smith1, Chen Yu1 1Department of Psychological and Brain Sciences, Indiana University, 2Department of Otolaryngology-Head and Neck Surgery, The Ohio State University, 3Department of Psychology, University of California, Riverside Young children do not passively observe the world, but rather actively explore and engage with their environment. This protocol provides guiding principles and practical recommendations for using head-mounted eye trackers to record infants' and toddlers' dynamic visual environments and visual attention in the context of natural behavior. Medicine Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury Allyson L. Huttinger1, Debra G. Wheeler1, Surya Gnyawali2, David Dornbos III1, Juliana M. Layzer3, Nicholas Venetos1, Spencer Talentino1, Nicholas J. Musgrave1, Cheyenne Jones1, Camille Bratton1, Matthew E. Joseph4, Chandan Sen2, Bruce A. Sullenger3, Shahid M. Nimjee1 1Department of Neurological Surgery, Ohio State University, 2Department of Surgery, Ohio State University, 3Department of Surgery, Duke University, 4Department of Internal Medicine, Ohio State University Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl3) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions. Biochemistry Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis Kang-Yi Su*1,2, Steven D. Goodman*3, Hung-Ming Lai1, Rong-Syuan Yen1, Wei-Yao Hu1, Wern-Cherng Cheng2, Liang-In Lin1,2, Ya-Chien Yang1,2, Woei-Horng Fang1,2 1Department of Clinical Laboratory Sciences and Medical Biotechnology, College of Medicine, National Taiwan University, 2Department of Laboratory Medicine, National Taiwan University Hospital, 3Center for Microbial Pathogenesis, Nationwide Children's Hospital and the Department of Pediatrics, The Ohio State University A non-labeled, non-radio-isotopic method for DNA polymerase proofreading and a DNA repair assay was developed by using high-resolution MALDI-TOF mass spectrometry and a single nucleotide extension strategy. The assay proved to be very specific, simple, rapid, and easy to perform for proofreading and repair patches shorter than 9-nucleotides. Medicine Collecting Hair Samples for Hair Cortisol Analysis in African Americans Kathy D. Wright1, Jodi L. Ford1, Joseph Perazzo2, Lenette M. Jones3, Sherrilynn Mahari4, Brent A. Sullenbarger5,6,7, Mark L. Laudenslager8 1College of Nursing, The Ohio State University, 2College of Nursing, University of Cincinnati, 3School of Nursing, University of Michigan, 4Licensed Cosmetologist Consultant, 5Clinical Research Manager, The Ohio State University, 6Biomedical Core Research Coordinator, The Ohio State University, 7College of Nursing Biomedical Core Research Laboratory, The Ohio State University, 8Department of Psychiatry, Behavioral Immunology & Endocrinology Laboratory, University of Colorado Denver Anschutz Medical Campus Hair cortisol concentration analysis provides an alternative to traditional measures of cortisol; however, to collect hair samples from African Americans, scientists need to be culturally informed and competent. The purpose of this protocol is to demonstrate a culturally informed technique to collect hair samples for cortisol analysis from African Americans. Neuroscience A Microbiomechanical System for Studying Varicosity Formation and Recovery in Central Neuron Axons Dustin Servello1, Yuanzheng Gu2,3, Chen Gu1,2 1Molecular, Cellular, and Developmental Biology Program, The Ohio State University, 2Department of Biological Chemistry and Pharmacology, The Ohio State University, 3Biogen This protocol describes a physiologically relevant, pressurized fluid approach for rapid and reversible induction of varicosities in neurons. Medicine Mechanical Ventilation Boot Camp Curriculum Jennifer Yee1,2, Alma Benner2, Jared Hammond2, Bethany Malone2, Charles Fuenning3, Richard George2,4, Rami A. Ahmed2,3,4 1Department of Emergency Medicine, The Ohio State University, 2Summa Health System, Akron City Hospital, 3Western Reserve Hospital, 4Northeast Ohio Medical University This curriculum outlines how to execute a simulation-based boot camp to teach providers how to manage mechanically ventilated patients. Environment The Calibration and Use of Capacitance Sensors to Monitor Stem Water Content in Trees Ashley M. Matheny1, Steven R. Garrity2, Gil Bohrer3 1Department of Geological Sciences, Jackson School of Geosciences, University of Texas at Austin, 2METER Group, Inc., USA, 3Department of Civil, Environmental and Geodetic Engineering, Ohio State University The hydraulic capacitance of biomass is a key component of the vegetation water budget, which serves as a buffer against short and long-term drought stresses. Here, we present a protocol for the calibration and use of soil moisture capacitance sensors to monitor water content in the stems of large trees. Biology Standardized Method for High-throughput Sterilization of Arabidopsis Seeds Benson E. Lindsey III*1, Luz Rivero*1, Chistopher S. Calhoun1, Erich Grotewold1,2, Jelena Brkljacic1 1Arabidopsis Biological Resource Center, Center for Applied Plant Sciences, The Ohio State University, 2Department of Molecular Genetics, Center for Applied Plant Sciences, The Ohio State University The objective of this study was to determine the effects of bleach and chlorine gas sterilization on seed germination of a range of Arabidopsis genotypes grown on sterile media. Optimized sterilization protocols have been developed to prevent the growth of microbial contaminants while providing satisfactory seed survival. 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. Chen1,5, Sanggu Kim6 1UCLA 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 Hatzakis1,2 1Department 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. Medicine Phosphorus-31 Magnetic Resonance Spectroscopy: A Tool for Measuring In Vivo Mitochondrial Oxidative Phosphorylation Capacity in Human Skeletal Muscle Vidhya Kumar1, Henry Chang1, David A. Reiter2, David P. Bradley3, Martha Belury4, Shana E. McCormack5, Subha V. Raman1 1Davis Heart and Lung Research Institute, The Ohio State University, 2Laboratory of Clinical Investigation, National Institute on Aging, 3Division of Endocrinology, Diabetes and Metabolism, The Ohio State University, 4Department of Human Sciences, Human Nutrition, The Ohio State University, 5Division of Endocrinology and Diabetes, Department of Pediatrics, University of Pennsylvania This work demonstrates the feasibility of an in vivo phosphorus-31 magnetic resonance spectroscopy (31PMRS) technique to quantify mitochondrial oxidative phosphorylation (OXPHOS) capacity in human skeletal muscle. Biochemistry Isolation of Cognate RNA-protein Complexes from Cells Using Oligonucleotide-directed Elution Gatikrushna Singh*1, Sarah M. Fritz*2, Arnaz Ranji2, Deepali Singh3, Kathleen Boris-Lawrie1,2 1Department of Veterinary & Biomedical Sciences, University of Minnesota, 2Department of Veterinary Biosciences, Ohio State University, 3School of Biotechnology, Gautam Buddha University This manuscript describes an approach to isolate select cognate RNPs formed in eukaryotic cells via a specific oligonucleotide-directed enrichment. We demonstrate the applicability of this approach by isolating a cognate RNP bound to the retroviral 5' untranslated region that is composed of DHX9/RNA helicase A. Medicine Drug Repurposing Hypothesis Generation Using the "RE:fine Drugs" System Kelly Regan1, Soheil Moosavinasab2, Philip Payne1, Simon Lin2 1Department of Biomedical Informatics, The Ohio State University, 2 Here we describe a protocol using the web-based drug repurposing hypothesis generation tool: "RE:fine Drugs." This protocol can be modified to a user's preferences at the level of the query type (gene, drug or disease) and/or the range of available advanced options. Environment Reliable Method for Assessing Seed Germination, Dormancy, and Mortality under Field Conditions Brian A. Pace1, Helen M. Alexander2, D. Jason Emry3, Kristin L. Mercer1 1Department of Horticulture and Crop Science, Ohio State University, 2Department of Ecology and Evolutionary Biology, University of Kansas, 3Department of Biology, Washburn University Here we present a protocol for assessing seed survivorship, germination and dormancy under field conditions using buried, labeled seed strips and tetrazolium chloride (TZ) viability testing. Behavior Electrophysiological Motor Unit Number Estimation (MUNE) Measuring Compound Muscle Action Potential (CMAP) in Mouse Hindlimb Muscles W. David Arnold1,2,3, Kajri A. Sheth1, Christopher G. Wier4, John T. Kissel1,3, Arthur H. Burghes1,3,4, Stephen J. Kolb1,3,4 1Department of Neurology, The Ohio State University Wexner Medical Center, 2Department of Physical Medicine and Rehabilitation, The Ohio State University, 3Department of Neuroscience, The Ohio State University Wexner Medical Center, 4Department of Biochemistry and Pharmacology, The Ohio State University Wexner Medical Center We present refined protocols that allow in vivo monitoring of motor unit function in the mouse. Techniques to measure compound muscle action potential (CMAP) and motor unit number estimation (MUNE) in the mouse hind limb muscles innervated by the sciatic nerve are described. Engineering Electron Channeling Contrast Imaging for Rapid III-V Heteroepitaxial Characterization Julia I. Deitz1, Santino D. Carnevale2, Steven A. Ringel3, David W. McComb3, Tyler J. Grassman1,2 1Department of Materials Science and Engineering, The Ohio State University, 2Department of Electrical and Computer Engineering, The Ohio State University, 3Institute of Materials Research, The Ohio State University The use of electron channeling contrast imaging in a scanning electron microscope to characterize defects in III-V/Si heteroexpitaxial thin films is described. This method yields similar results to plan-view transmission electron microscopy, but in significantly less time due to lack of required sample preparation. Chemistry Chemical Vapor Deposition of an Organic Magnet, Vanadium Tetracyanoethylene Megan Harberts1, Yu Lu2, Howard Yu1, Arthur J. Epstein1,2, Ezekiel Johnston-Halperin1 1Department of Physics, The Ohio State University, 2Department of Chemistry, The Ohio State University We present the synthesis of the organic-based ferrimagnet vanadium tetracyanoethylene (V[TCNE]x, x~2) via low temperature chemical vapor deposition (CVD). This optimized recipe yields an increase in Curie temperature from 400 K to over 600 K and a dramatic improvement in magnetic resonance properties. Medicine Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect Bo Zhang1,2, Jonathan P. Davis1, Mark T. Ziolo1 1Department of Physiology & Cell Biology, Davis Heart and Lung Research Institute, The Ohio State University, 2Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates. Behavior A Protocol for Housing Mice in an Enriched Environment Andrew M. Slater1, Lei Cao1 1Department of Molecular Virology, Immunology, and Medical Genetics, and Comprehensive Cancer Center, The Ohio State University Environmental enrichment for mice requires a complex and challenging setup, as well as comprehensive husbandry and handling techniques to assure robust metabolic and anti-cancer effects in the mice. This protocol provides detailed procedures to reproduce the above mentioned effects in mice. Medicine Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots Lu Xu1, Qiwen Shen1, Zhongqi Mao2, L. James Lee3, Ouliana Ziouzenkova1 1Department of Human Sciences, The Ohio State University, 2Department of Minimally Invasive Surgery, The First Affiliated Hospital of Soochow University, 3NSF Nanoscale Science and Engineering Center for Affordable Nanoengineering of Polymeric Biomedical Devices, The Ohio State University Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice. Medicine Method of Isolated Ex Vivo Lung Perfusion in a Rat Model: Lessons Learned from Developing a Rat EVLP Program Kevin Nelson1,2, Christopher Bobba1,2, Emre Eren3, Tyler Spata4, Malak Tadres2, Don Hayes, Jr.5,6, Sylvester M. Black3,7, Samir Ghadiali*1,2,3, Bryan A. Whitson*2,3,4 1Department of Biomedical Engineering, Ohio State University Wexner Medical Center, 2Davis Heart & Lung Research Institute, Ohio State University Wexner Medical Center, 3The Collaboration for Organ Perfusion, Protection, Engineering and Regeneration (COPPER) Laboratory, Ohio State University Wexner Medical Center, 4Division of Cardiac Surgery, Department of Surgery, Ohio State University Wexner Medical Center, 5Departments of Pediatrics and Internal Medicine, Ohio State University, 6 Ex-Vivo Lung Perfusion (EVLP) has allowed lung transplantation in humans to become more readily available by enabling the ability to assess organs and expand the donor pool. Here, we describe the development of a rat EVLP program and refinements that allow for a reproducible model for future expansion. Biology Quantitative Immunofluorescence Assay to Measure the Variation in Protein Levels at Centrosomes Shubhra Majumder1, Harold A. Fisk1 1Department of Molecular Genetics, The Ohio State University Here, a novel quantitative fluorescence assay is developed to measure changes in the level of a protein specifically at centrosomes by normalizing that protein’s fluorescence intensity to that of an appropriate internal standard. Medicine Neural Stem Cell Transplantation in Experimental Contusive Model of Spinal Cord Injury Stephana Carelli*1, Toniella Giallongo*1, Claudio Gerace1, Anthea De Angelis1, Michele D. Basso2, Anna Maria Di Giulio1, Alfredo Gorio1 1Laboratory of Pharmacology, Department of Health Sciences, University of Milan, 2Department of Neuroscience, College of Medicine, The Ohio State University Spinal cord injury is a traumatic condition that causes severe morbidity and high mortality. In this work we describe in detail a contusion model of spinal cord injury in mice followed by a transplantation of neural stem cells. Biology An Experimental and Bioinformatics Protocol for RNA-seq Analyses of Photoperiodic Diapause in the Asian Tiger Mosquito, Aedes albopictus Monica F. Poelchau1, Xin Huang1, Allison Goff1, Julie Reynolds2, Peter Armbruster1 1Department of Biology, Georgetown University, 2Insect Physiology Lab, EEOB, The Ohio State University RNA-Seq analyses are becoming increasingly important for identifying the molecular underpinnings of adaptive traits in non-model organisms. Here, a protocol to identify differentially expressed genes between diapause and non-diapause Aedes albopictus mosquitoes is described, from mosquito rearing, to RNA sequencing and bioinformatics analyses of RNA-Seq data. Biology Intravenous Injections in Neonatal Mice Sara E. Gombash Lampe1, Brian K. Kaspar2, Kevin D. Foust1 1Department of Neuroscience, Ohio State University, 2Center for Gene Therapy, Nationwide Children's Hospital Research Institute, Ohio State University Animal models of pediatric disease can experience early onset and aggressive disease progression. Clinically relevant therapy delivery to young mouse models can be technically difficult. This protocol describes a non-invasive intravenous injection method for newborn mice within the first two postnatal days of life. Bioengineering Bead Aggregation Assays for the Characterization of Putative Cell Adhesion Molecules Michelle R. Emond1, James D. Jontes1 1Department of Neuroscience, Ohio State University Here we present a simple, rapid method for characterizing the intrinsic adhesive properties of putative cell adhesion molecules. The secreted, epitope-tagged ectodomain of a cell adhesion molecule is captured from the culture medium on small, uniform functionalized beads. These beads can then be used immediately in simple bead aggregation assays. Medicine Collection and Extraction of Saliva DNA for Next Generation Sequencing Michael R. Goode1, Soo Yeon Cheong1, Ning Li1, William C. Ray1,2, Christopher W. Bartlett1,3 1 DNA extraction from saliva can provide a readily available source of high molecular weight DNA, with little to no degradation/fragmentation. This protocol provides optimized parameters for saliva collection/storage and DNA extraction to be of sufficient quality and quantity for downstream DNA assays with high quality requirements. Biology Isolation of Murine Valve Endothelial Cells Lindsey J. Miller1,2, Joy Lincoln2,3 1Molecular, Cellular and Developmental Biology Graduate Program, The Ohio State University, 2 The ability to isolate heart valve endothelial cells (VECs) is critical for understanding mechanisms of valve development, maintenance, and disease. Here we describe the isolation of VECs from embryonic and adult Tie2-GFP mice using FACS that will allow for studies determining the contribution of VECs in developmental and disease processes. Biology RNA Isolation from Mouse Pancreas: A Ribonuclease-rich Tissue Ana Clara P. Azevedo-Pouly1, Ola A. Elgamal1, Thomas D. Schmittgen1 1Pharmaceutics & Pharmaceutical Chemistry, The Ohio State University We report a procedure to isolate RNA with high integrity from the ribonuclease rich mouse pancreas. Biology Tissue Triage and Freezing for Models of Skeletal Muscle Disease Hui Meng1, Paul M.L. Janssen2, Robert W. Grange3, Lin Yang4, Alan H. Beggs5, Lindsay C. Swanson5, Stacy A. Cossette1,6, Alison Frase7, Martin K. Childers8, Henk Granzier9, Emanuela Gussoni5, Michael W. Lawlor1 1Division of Pediatric Pathology, Department of Pathology and Laboratory Medicine, Medical College of Wisconsin, 2Department of Physiology and Cell Biology, The Ohio State University, 3Department of Human Nutrition, Foods and Exercise, Virginia Tech, 4Division of Biomedical Informatics, Department of Biostatistics, Department of Computer Science, University of Kentucky, 5 The analysis of skeletal muscle tissues to determine structural, functional, and biochemical properties is greatly facilitated by appropriate preparation. This protocol describes appropriate methods to prepare skeletal muscle tissue for a broad range of phenotyping studies. Neuroscience Assaying DNA Damage in Hippocampal Neurons Using the Comet Assay Somaira Nowsheen1, Fen Xia2, Eddy S. Yang1,3 1Department 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. Lower1,2, Wei Lin3, Brian H. Lower2 1School 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 Zhao1,3 1Department 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. Hadley1 1Department 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. Villamena1,2 1The 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 Detloff1,2, Lesley C. Fisher1, Rochelle J. Deibert1, D. Michele Basso1 1Center 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 Lu2,3,4,5 1Center 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. Mangel1 1Department 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 Nakano1 1Department 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. Vyas1 1Department 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. Powell1,2, Tsonwin Hai1,2,3 1Molecular 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 Ransburgh1 1Department 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. Gnyawali3,4, Chandan K. Sen3 1Department 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. Gnyawali1,2, Sashwati Roy1,2, Jason Driggs1,2, Savita Khanna1,2, Thomas Ryan2,3, Chandan K. Sen2 1Department 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. Chiera1,2, John J. Finer1 1Department 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 Mittapalli1 1Department 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 Knaap1 1Department 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. Hadley1 1Department 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.
