Ben-Gurion University of the Negev 25 articles published in JoVE Biology Imaging Flow Cytometry to Study Microbial Autoaggregation Ronit Suissa1, Uzi Hadad2, Michael Meijler1, Ilana Kolodkin-Gal3,4 1Department of Chemistry, Ben-Gurion University of the Negev, 2Ilse Kats Institute for Nanoscale Science & Technology, Ben-Gurion University of the Negev, 3Department of Plant Pathology and Microbiology, Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, 4The Scojen institute for synthetic biology, Reichman university This protocol describes a quantitative approach to measure microbial autoaggregation using imaging flow cytometry. Bioengineering The Mechanics of (Poro-)Elastic Contractile Actomyosin Networks As a Model System of the Cell Cytoskeleton Sakshi Choudhary1, Gefen Livne1, Shachar Gat1, Anne Bernheim-Groswasser1,2 1Department of Chemical Engineering, Ben Gurion University of the Negev, 2Department of Chemical Engineering, Ilse Kats Institute for Nanoscale Science and Technology, Ben Gurion University of the Negev In this work, an in vitro reconstitution approach is employed to study the poroelasticity of actomyosin gels under controlled conditions. The dynamics of the actomyosin gel and the embedded solvent are quantified, through which the network poroelasticity is demonstrated. We also discuss the experimental challenges, common pitfalls, and relevance to cell cytoskeleton mechanics. Behavior Assessing Dominant-Submissive Behavior in Adult Rats Following Traumatic Brain Injury Dmitry Frank*1, Benjamin F. Gruenbaum*2, Michael Semyonov1, Yair Binyamin1, Olena Severynovska3, Ron Gal1, Amit Frenkel1, Boris Knazer4, Matthew Boyko1, Alexander Zlotnik1 1Division of Anesthesiology and Critical Care, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 2Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, 3Department of Biochemistry and Physiology of the Faculty of Biology and Ecology, Oles Gonchar of the Dnipro National University, 4Department of Ophthalmology, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev The present protocol describes a rat model of fluid percussion-induced traumatic brain injury followed by a series of behavioral tests to understand the development of dominant and submissive behavior. Using this model of traumatic brain injury in conjunction with specific behavioral tests enables the study of social impairments following brain injury. Biochemistry Motility of Single Molecules and Clusters of Bi-Directional Kinesin-5 Cin8 Purified from S. cerevisiae Cells Himanshu Pandey1, Tatiana Zvagelsky1, Mary Popov1, Mayan Sadan1, Neta Yanir1, Alina Goldstein-Levitin1, Nurit Siegler1, Shira Hershfinkel1, Yahel Abraham1, Roy Avraham1, Levi A. Gheber2, Larisa Gheber1 1Department of Chemistry, Ben-Gurion University of the Negev, Israel, 2Department of Biotechnology Engineering, Ben-Gurion University of the Negev, Israel The bi-directional mitotic kinesin-5 Cin8 accumulates in clusters that split and merge during their motility. Accumulation in clusters also changes the velocity and directionality of Cin8. Here, a protocol for motility assays with purified Cin8-GFP and analysis of motile properties of single molecules and clusters of Cin8 is described. Behavior A Complex Diving-For-Food Task to Investigate Social Organization and Interactions in Rats Benjamin F. Gruenbaum*1, Dmitry Frank*2, Shiri Savir2, Honore N. Shiyntum3, Ruslan Kuts2, Max Vinokur2, Israel Melamed4, Michael Dubilet2, Alexander Zlotnik2, Matthew Boyko2 1Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, 2Division of Anesthesiology and Critical Care, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 3 This protocol describes a method of examining social hierarchy in a rat model. Rats perform a complex diving-for-food task in which they form a distinct hierarchy according to their willingness to dive underwater and swim to obtain a food pellet. This method is used to understand decision making and social relationships among highly social animals in small groups. Neuroscience A Metric Test for Assessing Spatial Working Memory in Adult Rats Following Traumatic Brain Injury Dmitry Frank*1, Benjamin F. Gruenbaum*2, Israel Melamed3, Julia Grinshpun1, Yair Benjamin1, Ievgeni Vzhetson1, Nadia Kravchenko4, Michael Dubilet1, Matthew Boyko*1, Alexander Zlotnik*1 1Department of Anesthesiology and Critical Care, Soroka Medical Center, Ben-Gurion University of the Negev, 2Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, 3Department of Neurosurgery, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 4Department of Physiology, Faculty of Biology, Ecology and Medicine, Dnepropetrovsk State University Traumatic brain injury (TBI) is commonly associated with memory impairment. Here, we present a protocol to assess spatial working memory after TBI via a metric task. A metric test is a useful tool to study spatial working memory impairment after TBI. Neuroscience Measuring Post-Stroke Cerebral Edema, Infarct Zone and Blood-Brain Barrier Breakdown in a Single Set of Rodent Brain Samples Dmitry Frank*1, Benjamin F. Gruenbaum*2, Julia Grinshpun1, Israel Melamed3, Olena Severynovska4, Ruslan Kuts1, Michael Semyonov1, Evgeni Brotfain1, Alexander Zlotnik1, Matthew Boyko1 1Department of Anesthesiology and Critical Care, Soroka Medical Center, Ben-Gurion University of the Negev, 2Department of Anesthesiology, Yale University School of Medicine, 3Department of Neurosurgery, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 4Department of Physiology, Faculty of Biology, Ecology and Medicine, Dnepropetrovsk State University This protocol describes a novel technique of measuring the three most important parameters of ischemic brain injury on the same set of rodent brain samples. Using only one brain sample is highly advantageous in terms of ethical and economic costs. Neuroscience Laser-Induced Brain Injury in the Motor Cortex of Rats Ruslan Kuts*1, Israel Melamed*2, Honore N. Shiyntum3, Benjamin F. Gruenbaum4, Dmitry Frank1, Boris Knyazer5, Dmitry Natanel1, Olena Severynovska3, Max Vinokur1, Matthew Boyko1 1Division of Anesthesiology and Critical Care, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 2Department of Neurosurgery, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 3 The protocol presented here shows a technique to create a rodent model of brain injury. The method described here uses laser irradiation and targets motor cortex. Biology Using Caenorhabditis elegans to Screen for Tissue-Specific Chaperone Interactions Shiran Dror1, Tomer D. Meidan1, Ido Karady1, Anat Ben-Zvi1 1Department of Life Sciences, Ben-Gurion University of the Negev To study chaperone-chaperone and chaperone-substrate interactions, we perform synthetic interaction screens in Caenorhabditis elegans using RNA interference in combination with mild mutations or over-expression of chaperones and monitor tissue-specific protein dysfunction at the organismal level. Biology Fluorescence-Activated Cell Sorting for the Isolation of Scleractinian Cell Populations Grace A. Snyder1, William E. Browne2, Nikki Traylor-Knowles*1, Benyamin Rosental*3 1Department of Marine Biology and Ecology, Rosenstiel School of Marine and Atmospheric Science, University of Miami, 2Department of Biology, University of Miami, 3The Shraga Segal Department of Microbiology, Immunology, and Genetics, Faculty of Health Sciences, and Regenerative Medicine and Stem Cell Research Center, Ben Gurion University of the Negev Corals create biodiverse ecosystems important for both humans and marine organisms. However, we still do not understand the full potential and function of many coral cells. Here, we present a protocol developed for the isolation, labeling, and separation of stony coral cell populations. Neuroscience Induction of Diffuse Axonal Brain Injury in Rats Based on Rotational Acceleration Dmitry Frank*1, Israel Melamed*2, Benjamin F. Gruenbaum3, Julia Grinshpun1, Ruslan Kuts1, Rachel Shvartsur4, Abed N. Azab4, Mohamad H. Assadi5, Max Vinokur1, Matthew Boyko1 1Division of Anesthesia and Critical Care, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 2Department of Neurosurgery, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 3Department of Anesthesiology, Yale University School of Medicine, 4Recanati School for Community Health Professions, Faculty of Health Sciences, Ben-Gurion University of the Negev, 5Department of microbiology and immunology, Faculty of Health Sciences, Ben-Gurion University of the Negev This protocol validates a reliable, easy-to-perform and reproducible rodent model of brain diffuse axonal injury (DAI) that induces widespread white matter damage without skull fractures or contusions. Medicine Inducing Acute Liver Injury in Rats via Carbon Tetrachloride (CCl4) Exposure Through an Orogastric Tube Dmitry Frank*1, Shiri Savir*1, Benjamin F. Gruenbaum2, Israel Melamed3, Julia Grinshpun1, Ruslan Kuts1, Boris Knyazer4, Alexander Zlotnik1, Max Vinokur1, Matthew Boyko1 1Department of Anesthesiology and Critical Care, Soroka Medical Center, Ben-Gurion University of the Negev, 2Department of Anesthesiology, Yale University School of Medicine, 3Department of Neurosurgery, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev, 4Department of Ophthalmology, Soroka University Medical Center and the Faculty of Health Sciences, Ben-Gurion University of the Negev This protocol describes a common and feasible method of inducing acute liver injury (ALI) via CCl4 exposure through an orogastric tube. CCl4 exposure induces ALI through the formation of reactive oxygen species during its biotransformation in the liver. This method is used to analyze the pathophysiology of ALI and examine different hepatoprotective strategies. Developmental Biology Generation of a Human iPSC-Based Blood-Brain Barrier Chip Srikanth Jagadeesan1,2,3, Michael J. Workman4, Anna Herland5,6, Clive N. Svendsen4, Gad D. Vatine1,2,3 1The Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 2The Regenerative Medicine and Stem Cell (RMSC) Research Center, Ben-Gurion University of the Negev, 3The Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev, 4The Board of Governors Regenerative Medicine Institute, Cedars-Sinai Medical Center, 5Division of Micro and Nanosystems, KTH Royal Institute of Technology, 6AIMES, Department of Neuroscience, Karolinska Institutet The blood-brain barrier (BBB) is a multicellular neurovascular unit tightly regulating brain homeostasis. By combining human iPSCs and organ-on-chip technologies, we have generated a personalized BBB chip, suitable for disease modeling and CNS drug penetrability predictions. A detailed protocol is described for the generation and operation of the BBB chip. Cancer Research In Vitro Establishment of a Genetically Engineered Murine Head and Neck Cancer Cell Line using an Adeno-Associated Virus-Cas9 System Manu Prasad*1,2, Sankar Jagadeeshan*1,2, Maurizio Scaltriti3, Irit Allon2,4, Moshe Elkabets1,2 1The Shraga Segal Department of Microbiology, Immunology and Genetics, Ben-Gurion University of the Negev, 2Faculty of Health Sciences, Ben-Gurion University of the Negev, 3Human Oncology & Pathogenesis Program (HOPP), Memorial Sloan Kettering Cancer Center, 4Institute of Pathology, Barzilai University Medical Center Development of murine models with specific genes mutated in head and neck cancer patients is required for understanding of neoplasia. Here, we present a protocol for in vitro transformation of primary murine tongue cells using an adeno-associated virus-Cas9 system to generate murine HNC cell lines with specific genomic alterations. Neuroscience Wireless Electrophysiological Recording of Neurons by Movable Tetrodes in Freely Swimming Fish Lear Cohen*1,2, Ehud Vinepinsky*2,3, Ronen Segev1,2,3 1Department of Biomedical Engineering, Ben-Gurion University of the Negev, 2Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev, 3Department of Life Sciences, Ben-Gurion University of the Negev A novel wireless technique for recording extracellular neural signals from the brain of freely swimming goldfish is presented. The recording device is composed of two tetrodes, a microdrive, a neural data logger, and a waterproof case. All parts are custom-made except for the data logger and its connector. Biochemistry DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling Stefan Ilic1, Shira Cohen1, Ariel Afek2, Raluca Gordan2, David B. Lukatsky1, Barak Akabayov1 1Department of Chemistry, Ben-Gurion University of the Negev, 2Center for Genomic and Computational Biology, Department of Biostatistics and Bioinformatics, Duke University Protein binding microarray (PBM) experiments combined with biochemical assays link the binding and catalytic properties of DNA primase, an enzyme that synthesizes RNA primers on template DNA. This method, designated as high-throughput primase profiling (HTPP), can be used to reveal DNA-binding patterns of a variety of enzymes. Behavior A Middle Cerebral Artery Occlusion Technique for Inducing Post-stroke Depression in Rats Ruslan Kuts*1, Israel Melamed*2, Honore N. Shiyntum3, Dmitry Frank1, Julia Grinshpun1, Alexander Zlotnik1, Evgeni Brotfain1, Michael Dubilet1, Dmitry Natanel1, Matthew Boyko1 1Department of Anesthesiology and Critical Care, Soroka University Medical Center, Faculty of Health Sciences, Ben-Gurion University of the Negev, 2Department of Neurosurgery, Soroka University Medical Center, Faculty of Health Sciences, Ben-Gurion University of the Negev, 3 Here we present a protocol to induce post-stroke depression in rats by occluding the middle cerebral artery via the internal carotid artery. We use the Porsolt forced swim test and the sucrose preference test to confirm and evaluate induced depressive moods. Biochemistry A Colorimetric Method for Measuring Iron Content in Plants Jonas C. Gitz1,2, Noy Sadot1, Michele Zaccai1, Raz Zarivach1,2 1Department of Life Sciences, Ben-Gurion University of the Negev, 2National Institute for Biotechnology in the Negev (NIBN) We present a simple and reliable protocol for measuring iron content in plant tissues using the colorimetric Prussian Blue method. Behavior A New Method for Inducing a Depression-Like Behavior in Rats Vladimir Zeldetz*1, Dmitry Natanel*2, Matthew Boyko2, Alexander Zlotnik2, Honore N. Shiyntum3, Julia Grinshpun2, Dmitry Frank2, Ruslan Kuts2, Evgeni Brotfain2, Jochanan Peiser2 1Department of Emergent Medicine, Soroka University Medical Center, Ben-Gurion University of the Negev, 2Division of Anesthesiology and Critical Care, Soroka Medical Center, Ben-Gurion University of the Negev, 3 This protocol describes a new model by which healthy rats could contract depression over a given time periodthrough contagion by exposure to chronic unpredictable stressed (CUS) rats. Engineering High-speed Continuous-wave Stimulated Brillouin Scattering Spectrometer for Material Analysis Itay Remer1, Lear Cohen1, Alberto Bilenca1,2 1Biomedical Engineering Department, Ben-Gurion University of the Negev, 2Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev We describe the construction of a rapid continuous-wave-stimulated-Brillouin-scattering (CW-SBS) spectrometer. The spectrometer employs single-frequency diode-lasers and an atomic vapor notch-filter to acquire transmission spectra of turbid/non-turbid samples with high spectral-resolution at speeds up to 100-fold faster than those of existing CW-SBS spectrometers. This improvement enables high-speed Brillouin material analysis. Neuroscience Assay to Measure Nucleocytoplasmic Transport in Real Time within Motor Neuron-like NSC-34 Cells Tom Shani1, Moshe Levy1, Adrian Israelson1,2 1Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 2The Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev This protocol describes a method to sensitively measure the nucleocytoplasmic transport rate within motor neuron-like NSC-34 cells by quantifying the real-time change in the nuclear import of a NLS-NES-GFP protein. Behavior How to Use the H1 Deep Transcranial Magnetic Stimulation Coil for Conditions Other than Depression Aron Tendler1, Yiftach Roth1, Noam Barnea-Ygael2, Abraham Zangen2 1Brainsway LTD, 2Department of Life Sciences, Ben Gurion University of the Negev The H1 deep transcranial magnetic stimulation coil is FDA-cleared for the treatment of depression. We demonstrate how to utilize the H1 for other conditions, such as auditory hallucinations and PTSD, by moving the helmet to different locations over the subject's skull. Bioengineering Three-Dimensionally Printed Microfluidic Cross-flow System for Ultrafiltration/Nanofiltration Membrane Performance Testing Nathaniel C. Wardrip1, Christopher J. Arnusch1 1Department of Desalination and Water Treatment, Zuckerberg Institute for Water Research, Jacob Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Sede-Boqer Campus Design and fabrication of a three-dimensionally (3-D) printed microfluidic cross-flow filtration system is demonstrated. The system is used to test performance and observe fouling of ultrafiltration and nanofiltration (thin film composite) membranes. Neuroscience Telomerase Activity in the Various Regions of Mouse Brain: Non-Radioactive Telomerase Repeat Amplification Protocol (TRAP) Assay Yossi Grin1, Tamar Admoni1, Esther Priel1 1The Shraga Segal Department of Immunology, Microbiology & Genetics, Faculty of Health Sciences, Ben-Gurion University of the Negev Telomerase is expressed in the neonatal brain and also in distinct regions of the adult brain. We present a non-toxic time saving TRAP assay for the analysis of telomerase activity in various regions of the mouse brain and detection of differences in telomerase activity between male and female mouse brains. Biology Using Caenorhabditis elegans as a Model System to Study Protein Homeostasis in a Multicellular Organism Ido Karady1, Anna Frumkin1, Shiran Dror1, Netta Shemesh1, Nadav Shai1, Anat Ben-Zvi1 1Department of Life Sciences, National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev To study the relationship between protein homeostasis, stress and aging, we monitored changes in protein folding by following protein dysfunction, protein localization in the cell and protein stability at the organismal, cellular and protein levels, using the genetically tractable metazoan Caenorhabditis elegans as a model system.
