University of Bern View Institution's Website 40 articles published in JoVE Bioengineering Use of the MicroSiM (µSiM) Barrier Tissue Platform for Modeling the Blood-Brain Barrier Molly C. McCloskey1, Pelin Kasap2, Michelle Trempel1, Louis P. Widom3, Julia Kuebel1, Kaihua Chen1, Thomas R. Gaborski3, Britta Engelhardt2, James L. McGrath1 1Department of Biomedical Engineering, University of Rochester, 2Theodor Kocher Institute, University of Bern, 3Department of Biomedical Engineering, Rochester Institute of Technology This report provides protocols for assembly, cell culture, and assays on the µSiM platform for the construction of blood-brain barrier models. Cancer Research Establishment of Pancreatic Cancer-Derived Tumor Organoids and Fibroblasts From Fresh Tissue Jesús Frutos Díaz-Alejo*1,2,3,4, Simon April-Monn*5, Marina Cihova6, Verona Buocikova6, Jorge Villalón López1,3, Maria Urbanova6, Carmen G. Lechuga7, Miroslav Tomas6,8, Peter Dubovan6,8, Bárbara Luna Sánchez3, Sonia Camaño Páez3, Alfonso Sanjuanbenito2,9, Eduardo Lobo9, Estefanía Romio de la Heras10, Carmen Guerra2,7, Carolina de la Pinta11, Emma Barreto Melian1,2, Mercedes Rodríguez Garrote1,2, Alfredo Carrato1,2,4, Laura Ruiz-Cañas3,12,13, Bruno Sainz, Jr.2,3,12,13, Ana Torres3, Bozena Smolkova6, Julie Earl1,2,3 1Molecular Epidemiology and Predictive Tumor Markers Group, Area 3, Ramón y Cajal Health Research Institute (IRYCIS), 2The Biomedical Research Network in Cancer (CIBERONC), 3Biobank and Biomodels Platform (PT20/0045), ISCIII research and development platforms in biomedicine and health sciences, BioBank Hospital Ramón y Cajal-IRYCIS, Spanish National Biobanks Network (ISCIII Biobank Register No. B.0000678), Ramón y Cajal Health Research Institute (IRYCIS), 4Faculty of Medicine, University of Alcalá de Henares, 5Institute of Tissue Medicine and Pathology, University of Bern, 6Department of Molecular Oncology, Cancer Research Institute, Biomedical Research Center of the Slovak Academy of Sciences, 7Experimental Oncology, Molecular Oncology Program, Centro Nacional de Investigaciones Oncológicas (CNIO), 8Department of Surgical Oncology, National Cancer Institute, Slovak Medical University, 9Pancreatic and Biliopancreatic Surgery Unit, Hospital Universitario Ramón y Cajal, 10Department of Pathology, Hospital Universitario Ramón y Cajal, 11Department of Radiation Oncology, Hospital Universitario Ramón y Cajal, 12Department of Cancer, Instituto de Investigaciones Biomédicas “Alberto Sols” (IIBM), 13Cancer Stem Cell and Fibroinflammatory Group, Chronic Diseases and Cancer, Area 3, IRYCIS Tumor organoids have revolutionized cancer research and the approach to personalized medicine. They represent a clinically relevant tumor model that allows researchers to stay one step ahead of the tumor in the clinic. This protocol establishes tumor organoids from fresh pancreatic tumor tissue samples and patient-derived xenografts of pancreatic adenocarcinoma origin. Bioengineering Differentiation of Human Induced Pluripotent Stem Cells to Brain Microvascular Endothelial Cell-Like Cells with a Mature Immune Phenotype Kinya Matsuo1, Britta Engelhardt2, Hideaki Nishihara3 1Graduate School of Medicine, Yamaguchi University, 2Theodor Kocher Institute, University of Bern, 3Department of Neurotherapeutics, Yamaguchi University of Medicine Here, we describe a protocol, the extended endothelial cell culture method (EECM), that allows differentiation of pluripotent stem cells to brain microvascular endothelial cell (BMEC)-like cells. These cells show endothelial cell adhesion molecule expression and are thus a human blood-brain barrier model suitable to study immune cell interactions in vitro. Neuroscience Measuring the Influence of Magnetic Vestibular Stimulation on Nystagmus, Self-Motion Perception, and Cognitive Performance in a 7T MRT Gerda Wyssen1,6, Miranda Morrison2, Athanasia Korda2, Wilhelm Wimmer2,3, Jorge Otero-Millan4, Matthias Ertl1, Andreas A. Szukics1,6, Thomas Wyss2, Franca Wagner5,6, Marco D. Caversaccio2,3, Georgios Mantokoudis2,6, Fred W. Mast1,6 1Department of Psychology, University of Bern, 2Department of Otorhinolaryngology, Head and Neck Surgery, lnselspital, University Hospital Bern and University of Bern, 3Hearing Research Laboratory, ARTORG Center for Biomedical Engineering Research, University of Bern, 4Herbert Wertheim School of Optometry and Vision Science, University of California, 5University Department of Diagnostic and Interventional Neuroradiology, Inselspital, Bern University Hospital, University of Bern, 6Translational Imaging Center (TIC), Swiss Institute for Translational and Entrepreneurial Medicine In this article, we describe the experimental setup, material, and procedures to assess reflexive eye movements, self-motion perception, and cognitive tasks under magnetic vestibular stimulation, as well as the anatomical orientation of the vestibular organs, in a 7 Tesla Magnetic resonance tomography (7T-MRT) scanner. Behavior Estimating Vestibular Perceptual Thresholds Using a Six-Degree-Of-Freedom Motion Platform Matthias Ertl1, Daniel C. Fitze1, Gerda Wyssen1, Fred W. Mast1 1Institute of Psychology, University of Bern In this article, we describe the methods, procedures, and technologies required to estimate vestibular perceptual thresholds using a six-degree-of-freedom motion platform. Medicine Robotic Cochlear Implantation for Direct Cochlear Access Marco Caversaccio1,2, Georgios Mantokoudis1, Franca Wagner3, Philipp Aebischer1,2, Stefan Weder1,2, Wilhelm Wimmer1,2 1Department of Otorhinolaryngology, Head and Neck Surgery, Inselspital, Bern University Hospital, University of Bern, 2Hearing Research Laboratory, ARTORG Center for Biomedical Engineering Research, University of Bern, 3Department of Diagnostic and Interventional Neuroradiology, Inselspital, Bern University Hospital Robotic cochlear implantation is a procedure for minimally invasive inner ear access. Compared to conventional surgery, robotic cochlear implantation involves additional steps that need to be carried out in the operating room. In this article we give a description of the procedure and highlight the important aspects of robotic cochlear implantation. Neuroscience Using a Cell-Tracer Injection to Investigate the Origin of Neointima-Forming Cells in a Rat Saccular Side Wall Model Stefan Wanderer*1,2, Basil E. Grüter*1,2, Jeannine Kümin1,2, Gwendoline Boillat1,2, Sivani Sivanrupan2, Kristina Catalano1,2, Michael von Gunten3, Hans Rudolf Widmer4, Serge Marbacher1,2,5, Lukas Andereggen1,2,5 1Department of Neurosurgery, Kantonsspital Aarau, 2Cerebrovascular Research Group, Department for BioMedical Research, University of Bern, 3Institute of Pathology Laenggasse, 4Department of Neurosurgery, Neurocenter and Regenerative Neuroscience Cluster, Inselspital, Bern University Hospital, University of Bern, 5Faculty of Medicine, University of Bern We performed a one-point, lipophilic cell-tracer injection to track endothelial cells, followed by an arteriotomy and suturing of sidewall aneurysms on the abdominal rat aorta. Neointima formation seemed dependent on the parent artery in decellularized aneurysms and was promoted by the recruitment from aneurysm wall cells in vital cell-rich walls. Medicine Performing Intracochlear Electrocochleography During Cochlear Implantation Klaus Schuerch1,2, Manuel Waser1, Georgios Mantokoudis1, Lukas Anschuetz1, Wilhelm Wimmer1,2, Marco Caversaccio1,2, Stefan Weder1 1Department of Otorhinolaryngology, Head and Neck Surgery, Inselspital, Bern University Hospital, University of Bern, 2Hearing Research Laboratory, ARTORG Center for Biomedical Engineering Research, University of Bern Electrocochleography (ECochG) measures inner ear potentials generated in response to acoustic stimulation. In cochlear implant (CI) candidates, such inner ear potentials can be measured directly with the implant electrodes. In this video, we systematically explain how to perform ECochG recordings during CI surgery. Medicine The Microscopic Transcanal Approach in Stapes Surgery Revisited Georgios Mantokoudis1, Stefan Weder1, Lukas Anschuetz1, Sandra Gunser1, Urs Borner1, Marco D. Caversaccio1 1Department of Otorhinolaryngology, Head and Neck Surgery, lnselspital, Bern University Hospital, University of Bern This article describes the microscopic transcanal technique for stapes surgery, providing step-by-step surgical instructions for familiarizing surgeons with this approach. Medicine Multiphoton Intravital Imaging for Monitoring Leukocyte Recruitment during Arteriogenesis in a Murine Hindlimb Model Manuel Lasch1,2,3, Mykhailo Vladymyrov4, Dominic van den Heuvel1,5, Philipp Götz1,3, Elisabeth Deindl1,3, Hellen Ishikawa-Ankerhold1,5 1Walter-Brendel-Centre of Experimental Medicine, University Hospital, Ludwig-Maximilians-Universität München, 2Department of Otorhinolaryngology, Head & Neck Surgery, University Hospital, Ludwig-Maximilians-Universität München, 3Biomedical Center, Institute of Cardiovascular Physiology and Pathophysiology, Faculty of Medicine, Ludwig-Maximilians-Universität München, 4TKI, University of Bern, 5Department of Internal Medicine I and Cardiology, University Hospital, Ludwig-Maximilians-Universität München The recruitment of leukocytes and platelets constitutes an essential component necessary for the effective growth of collateral arteries during arteriogenesis. Multiphoton microscopy is an efficient tool for tracking cell dynamics with high spatio-temporal resolution in vivo and less photo-toxicity to study leukocyte recruitment and extravasation during arteriogenesis. Biology 5-Ethynyl-2'-Deoxyuridine/Phospho-Histone H3 Dual-Labeling Protocol for Cell Cycle Progression Analysis in Drosophila Neural Stem Cells Rohan Chippalkatti1,2, Beat Suter1 1Cell Biology, University of Bern, 2Department of life sciences and medicine, University of Luxembourg Cell cycle analysis with 5-ethynyl-2'-deoxyuridine (EdU) and phospho-histone H3 (pH3) labeling is a multi-step procedure that may require extensive optimization. Here, we present a detailed protocol that describes all steps for this procedure including image analysis and quantification to distinguish cells in different cell cycle phases. Medicine Creation of Two Saccular Elastase-Digested Aneurysms with Different Hemodynamics in One Rabbit Gwendoline Boillat1,2, Tim Franssen2, Basil Grüter1,2, Stefan Wanderer1,2, Kristina Catalano2, Daniela Casoni3, Lukas Andereggen1,2, Serge Marbacher1,2 1Department of Neurosurgery, Kantonsspital Aarau, 2Cerebrovascular Research Group, Department for BioMedical Research, University of Bern, 3Experimental Surgery Facility, Department for Biomedical Research, Faculty of Medicine, University of Bern This protocol describes the steps for the creation of a rabbit model with two elastase-digested aneurysms with different hemodynamics (stump and bifurcation constellation). This allows the testing of novel endovascular devices in aneurysms with different angioarchitecture and hemodynamic conditions within a single animal. Neuroscience Arterial Pouch Microsurgical Bifurcation Aneurysm Model in the Rabbit Stefan Wanderer1,2, Claudia Waltenspuel2, Basil E. Grüter1,2, Fabio Strange1,2, Sivani Sivanrupan2, Luca Remonda3, Hans Rudolf Widmer4, Daniela Casoni5, Lukas Andereggen1,2, Javier Fandino1,2, Serge Marbacher1,2 1Department of Neurosurgery, Kantonsspital Aarau, 2Cerebrovascular Research Group, Department for BioMedical Research, University of Bern, 3Division of Neuroradiology, Department of Radiology, Kantonsspital Aarau, 4Department of Neurosurgery, Neurocenter and Regenerative Neuroscience Cluster, Inselspital, Bern University Hospital, University of Bern, 5Department for Biomedical Research, Faculty of Medicine, University of Bern Developing and testing endovascular devices for intracranial aneurysm treatment is still of great importance. Most aneurysm models used today miss either the important characteristics of an arterial degenerated wall or the hemodynamics of a true bifurcation. Therefore, we aimed to design a novel arterial pouch bifurcation model in rabbits. Neuroscience Muscle Velocity Recovery Cycles to Examine Muscle Membrane Properties Agnes Witt1, Hugh Bostock2, Werner J. Z'Graggen3, S. Veronica Tan4, Alexander Gramm Kristensen1, Rikke Søgaard Kristensen1, Lotte Hardbo Larsen1, Zennia Zeppelin1, Hatice Tankisi1 1Department of Clinical Neurophysiology, Aarhus University Hospital, 2UCL Queen Square Institute of Neurology, Queen Square House, 3Departments of Neurology and Neurosurgery, Inselspital, Bern University Hospital, University of Bern, 4MRC Centre for Neuromuscular Diseases, UCL Institute of Neurology, The National Hospital for Neurology and Neurosurgery Presented here is a protocol for the recording of muscle velocity recovery cycles (MVRCs), a new method of examining muscle membrane properties. MVRCs enable in vivo assessment of muscle membrane potential and alterations in muscle ion channel function in relation to pathology, and it enables the demonstration of muscle depolarization in neurogenic muscles. Neuroscience Fluorescence Angiography for Evaluation of Aneurysm Perfusion and Parent Artery Patency in Rat and Rabbit Aneurysm Models Fabio Strange1,2, Sivani Sivanrupan2, Basil E. Gruter1,2, Jeannine Rey2, Dominik Taeschler2, Javier Fandino1,2, Serge Marbacher1,2 1Department of Neurosurgery, Kantonsspital Aarau, 2Cerebrovascular Research Group, Department for BioMedical Research (DBMR), University of Bern We present a protocol to efficiently evaluate aneurysm perfusion and vessel patency of sidewall aneurysm in rats and rabbits, using fluorescein-based fluorescence video angiography (FVA). With a positive predictive value of 92.6%, it is a simple but very effective and economical method with no special equipment required. Neuroscience Targeting Alpha Synuclein Aggregates in Cutaneous Peripheral Nerve Fibers by Free-floating Immunofluorescence Assay Elena Vacchi1, Sandra Pinton1, Alain Kaelin-Lang1,2,3,4, Giorgia Melli1,2 1Laboratory for Biomedical Neurosciences, Neurocenter of Southern Switzerland, 2Neurology Department, Neurocenter of Southern Switzerland, 3Department of Neurology, Inselspital, Bern University Hospital, University of Bern, 4Faculty of Biomedical Sciences, Università della Svizzera Italiana Here, we present a protocol for a free-floating indirect immunofluorescence assay on skin biopsy sections that allows for the identification of disease specific conformation variants of alpha synuclein involved in Parkinson disease and multiple proteins of the peripheral nervous system. Immunology and Infection Visualizing Impairment of the Endothelial and Glial Barriers of the Neurovascular Unit during Experimental Autoimmune Encephalomyelitis In Vivo Silvia M. Tietz1, Britta Engelhardt1 1Theodor Kocher Institute, University of Bern Here, we present protocols to investigate impairment of the neurovascular unit during experimental autoimmune encephalomyelitis in vivo. We specifically address how to determine blood-brain barrier permeability and gelatinase activity involved in leukocyte migration across the glia limitans. Environment Detection of Tilapia Lake Virus Using Conventional RT-PCR and SYBR Green RT-qPCR Pamela Nicholson1, Pattarasuda Rawiwan2, Win Surachetpong2 1Institute of Veterinary Bacteriology Vetsuisse Faculty, University of Bern, 2Department of Veterinary Microbiology and Immunology and Center for Advanced Studies for Agriculture and Food, Kasetsart University Institute for Advanced Studies, Kasetsart University This protocol diagnoses Tilapia Lake Virus (TiLV) in tilapia tissues using RT-PCR methodologies. The entire method is described from tissue dissection to total RNA extraction, followed by cDNA synthesis and detection of TiLV by either conventional PCR or quantitative PCR using dsDNA binding a fluorescent binding dye. Immunology and Infection Imaging Cell Interaction in Tracheal Mucosa During Influenza Virus Infection Using Two-photon Intravital Microscopy Miguel Palomino-Segura*1,2, Tommaso Virgilio*1,2, Diego Morone1, Diego U. Pizzagalli1,3, Santiago F. Gonzalez1 1Faculty of Biomedical Sciences, Institute for Research in Biomedicine, Università della Svizzera italiana (USI), 2Graduate School of Cellular and Molecular Sciences, Faculty of Medicine, University of Bern, 3Institute of Computational Science, Università della Svizzera italiana (USI) In this study, we present a protocol to perform two-photon intravital imaging and cell interaction analysis in the murine tracheal mucosa after infection with influenza virus. This protocol will be relevant for researchers studying immune cell dynamics during respiratory infections. Immunology and Infection Preparation of Murine Submandibular Salivary Gland for Upright Intravital Microscopy Xenia Ficht*1, Flavian Thelen*1, Bettina Stolp*1,2, Jens V. Stein1 1Theodor-Kocher Institute, University of Bern, 2Center for Infectious Diseases, Integrative Virology, University Clinic of Heidelberg We describe a protocol to surgically expose and stabilize the murine submandibular salivary gland for intravital imaging using upright intravital microscopy. This protocol is easily adaptable to other exocrine glands of the head and neck region of mice and other small rodents. Environment On the Preparation and Testing of Fuel Cell Catalysts Using the Thin Film Rotating Disk Electrode Method Masanori Inaba1, Jonathan Quinson1, Jan Rudolf Bucher2, Matthias Arenz1,2 1Nano-Science Center, Department of Chemistry, University of Copenhagen, 2Department of Chemistry and Biochemistry, University of Bern Preparing and testing Pt/C fuel cell catalysts is subject to continuous discussion in the scientific community with respect to reproducibility and best practice. With the presented work, we intend to present a step-by-step tutorial to make and test Pt/C catalysts, which can serve as benchmark for novel catalyst systems. Medicine Improvement of a Closed Chest Porcine Myocardial Infarction Model by Standardization of Tissue and Blood Sampling Procedures Mai M. Abdelhafez1,2, Jane Shaw1, Jonas Wilbs3, Alain Despont1, Robert Rieben1 1Department for Biomedical Research, University of Bern, 2Graduate School for Cellular and Biomedical Sciences, University of Bern, 3Institute of Chemical Sciences and Engineering, Ecole Polytechnique Fédérale de Lausanne (EPFL) Here we demonstrate a protocol to standardize sampling procedures of an established porcine model of acute myocardial infarction in order to increase its translational value in the understanding of the pathophysiology of myocardial ischemia/reperfusion injury and to test novel drug candidates. Neuroscience Müller Glia Cell Activation in a Laser-induced Retinal Degeneration and Regeneration Model in Zebrafish Federica M. Conedera1,2,3, Petra Arendt1, Carolyn Trepp1,2,3, Markus Tschopp1, Volker Enzmann1,2 1Department of Ophthalmology, University Hospital of Bern, University of Bern, 2Department of Clinical Research, University of Bern, 3Graduate School for Cellular and Biomedical Sciences, University of Bern The zebrafish is a popular animal model to study mechanisms of retinal degeneration/regeneration in vertebrates. This protocol describes a method to induce localized injury disrupting the outer retina with minimal damage to the inner retina. Subsequently, we monitor in vivo the retinal morphology and the Müller glia response throughout retinal regeneration. Medicine Assessment of the Anticoagulant and Anti-inflammatory Properties of Endothelial Cells Using 3D Cell Culture and Non-anticoagulated Whole Blood Riccardo Sfriso1,2, Anjan Bongoni3, Yara Banz4, Nikolai Klymiuk5, Eckhard Wolf5, Robert Rieben1 1Department of Clinical Research, University of Bern, 2Graduate School for Cellular and Biomedical Sciences, University of Bern, 3 We present an in vitro model which allows the study and analysis of coagulation in whole, non-anticoagulated blood. Anticoagulation in the system depends on the natural anticoagulation effect of healthy endothelial cells and endothelial cell activation will result in clotting. Biology High-resolution Respirometry to Assess Mitochondrial Function in Permeabilized and Intact Cells Siamak Djafarzadeh1, Stephan M. Jakob1 1Department of Intensive Care Medicine, Inselspital, Bern University Hospital, University of Bern High-resolution respirometry is used to determine mitochondrial oxygen consumption. This is a straightforward technique to determine mitochondrial respiratory chain complexes' (I-IV) respiratory rates, maximal mitochondrial electron transport system capacity, and mitochondrial outer membrane integrity. Biology Xenopus Oocytes: Optimized Methods for Microinjection, Removal of Follicular Cell Layers, and Fast Solution Changes in Electrophysiological Experiments Maria C. Maldifassi*1, Nisa Wongsamitkul*1, Roland Baur1, Erwin Sigel1 1Institute of Biochemistry and Molecular Medicine, University of Bern Optimized procedures for the isolation of single follicles, cytoplasmic RNA microinjections, the removal of surrounding cell layers, and protein expression in Xenopus oocytes are described. In addition, a simple method for fast solution changes in electrophysiological experiments with ligand-gated ion channels is presented. Neuroscience Spiral Ganglion Neuron Explant Culture and Electrophysiology on Multi Electrode Arrays Stefan Hahnewald1,2, Marta Roccio1,2, Anne Tscherter3, Jürg Streit3, Ranjeeta Ambett1,2, Pascal Senn1,2,4 1Department of Otorhinolaryngology, Inselspital and University of Bern, 2Department of Clinical Research, Inselspital and University of Bern, 3Department of Physiology, University of Bern, 4Department of Clinical Neurosciences, Service ORL & HNS, University Hospital Geneva We present a protocol to culture primary murine spiral ganglion neuron explants on multi electrode arrays to study neuronal response profiles and optimize stimulation parameters. Such studies aim to improve the neuron-electrode interface of cochlear implants to benefit hearing in patients as well as the energy consumption of the device. Behavior Concurrent Electroencephalography Recording During Transcranial Alternating Current Stimulation (tACS) Kristoffer D. Fehér1, Yosuke Morishima1,2 1Division of Systems Neuroscience of Psychopathology, Translational Research Center, University Hospital of Psychiatry, University of Bern, 2Japan Science and Technology Agency, PRESTO In this article we explain how to set up a concurrent transcranial alternating current stimulation and EEG experiment. Neuroscience Investigating Functional Regeneration in Organotypic Spinal Cord Co-cultures Grown on Multi-electrode Arrays Martina Heidemann1, Jürg Streit1, Anne Tscherter1 1Department of Physiology, University of Bern Here we present a protocol that is based on spinal cord slices cultured on multi-electrode arrays to study functional regeneration of propriospinal connections in vitro. Biology Bone Conditioned Medium: Preparation and Bioassay Jordi Caballé-Serrano1,2,3, Kosaku Sawada2,5, Guenther Schuldt Filho1,2,6, Dieter D. Bosshardt1,4, Daniel Buser1, Reinhard Gruber1,2 1Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern, 2Laboratory of Oral Cell Biology, School of Dental Medicine, University of Bern, 3Department of Oral and Maxillofacial Surgery, School of Dental Medicine, Universitat Internacional de Catalunya, 4Robert K. Schenk Laboratory of Oral Histology, School of Dental Medicine, University of Bern, 5Department of Cranio Maxillofacial Surgery, Inselspital, University of Bern, 6Department of Implant Dentistry, School of Dentistry, Universidade Federal de Santa Catarina We describe here how to prepare bone-conditioned medium (BCM) and test its activity in vitro. Biology Methylnitrosourea (MNU)-induced Retinal Degeneration and Regeneration in the Zebrafish: Histological and Functional Characteristics Ellinor Maurer1, Markus Tschopp1,2, Christoph Tappeiner1, Pauline Sallin3, Anna Jazwinska3, Volker Enzmann1 1Department of Ophthalmology, Inselspital, University of Bern, 2Department of Ophthalmology, University Hospital of Basel, 3Department of Biology, University of Fribourg Herein we demonstrate quantification of retinal de- and regeneration and its impact on visual function using N-methyl-N-nitrosourea in the adult zebrafish. Loss of visual acuity and decreased photoreceptor numbers were followed by proliferation in the inner nuclear layer. Complete morphological and functional regeneration occurred 30 days after the initial treatment. Medicine The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects Lukas Andereggen3,4,5, Volker Neuschmelting1,6, Michael von Gunten7, Hans Rudolf Widmer5, Jukka Takala1, Stephan M. Jakob1, Javier Fandino1,2, Serge Marbacher1,2 1Department of Intensive Care Medicine, University and Bern University Hospital (Inselspital), 2Department of Neurosurgery, Kantonsspital Aarau, 3 The experimental intracranial pressure-controlled blood shunt subarachnoid hemorrhage (SAH) model in the rabbit combines the standard procedures — subclavian artery cannulation and transcutaneous cisterna magna puncture, which enables close mimicking of human pathophysiological conditions after SAH. We present step-by-step instructions and discuss key surgical points for successful experimental SAH creation. Medicine A Next-generation Tissue Microarray (ngTMA) Protocol for Biomarker Studies Inti Zlobec1, Guido Suter1, Aurel Perren1, Alessandro Lugli1 1Institute of Pathology, University of Bern The protocol aims at optimizing the construction and quality of tissue microarrays for biomarker research. It includes aspects of planning and design, digital pathology, virtual slide annotation, and automated tissue arraying. Chemistry Nucleoside Triphosphates - From Synthesis to Biochemical Characterization Marcel Hollenstein1, Christine Catherine Smith1, Michael Räz1 1Department of Chemistry and Biochemistry, University of Bern The protocol described herein aims to explain and abridge the numerous obstacles in the way of the intricate route leading to modified nucleoside triphosphates. Consequently, this protocol facilitates both the synthesis of these activated building-blocks and their availability for practical applications. Immunology and Infection Isolation of Myeloid Dendritic Cells and Epithelial Cells from Human Thymus Christina Stoeckle1,2, Ioanna A. Rota1, Eva Tolosa3, Christoph Haller4, Arthur Melms5, Eleni Adamopoulou1 1Department of General Neurology, Hertie Institute for Clinical Brain Research, 2Institute of Pharmacology, University of Bern, 3Department of Immunology, University Medical Center Hamburg-Eppendorf, 4Department of Thoracic and Cardiovascular Surgery, University Clinic Tuebingen, 5Department of Neurology, University Hospital Erlangen This protocol details a method to isolate antigen presenting cells from human thymus via different steps of enzymatic digestion of the tissue followed by density centrifugation of the single cell suspension and finally magnetic and/or FACS sorting of the cell populations of interest. Medicine Bioluminescent Orthotopic Model of Pancreatic Cancer Progression Ming G. Chai1, Corina Kim-Fuchs1,2, Eliane Angst2, Erica K. Sloan1,3 1Monash Institute of Pharmaceutical Sciences, Monash University, 2Department of Visceral Surgery and Medicine, University of Bern, 3Cousins Center for Neuroimmunology, University of California Los Angeles Improved understanding of pancreatic cancer biology is critically needed to enable the development of better therapeutic options to treat pancreatic cancer. To address this need, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression using in vivo bioluminescence imaging. Bioengineering Determination of the Transport Rate of Xenobiotics and Nanomaterials Across the Placenta using the ex vivo Human Placental Perfusion Model Stefanie Grafmüller1,2,3, Pius Manser2, Harald F. Krug2, Peter Wick2, Ursula von Mandach1 1Department of Obstetrics, Perinatal Pharmacology, University Hospital Zurich, 2Laboratory for Materials - Biology Interactions, EMPA Swiss Federal Laboratories for Materials Testing and Research, 3Graduate School for Cellular and Biomedical Sciences, University of Bern The ex vivo dual recirculating human placental perfusion model can be used to investigate the transfer of xenobiotics and nanoparticles across the human placenta. In this video protocol we describe the equipment and techniques required for a successful execution of a placenta perfusion. Immunology and Infection Diagnosing Pulmonary Tuberculosis with the Xpert MTB/RIF Test Thomas Bodmer1, Angelika Ströhle2 1Institute for Infectious Diseases, University of Bern, 2MCL Laboratories Inc. The Xpert MTB/RIF test integrates sample decontamination, hands-free operation, on-board sample processing, and ultra-sensitive hemi-nested PCR for the simultaneous detection of Mycobacterium tuberculosis and rifampicin resistance, either in expectorated sputum or concentrated sputum sediments, in approximately two hours. Testing is standardized and requires only moderate laboratory infrastructure and training. Bioengineering Preparation of Intact Bovine Tail Intervertebral Discs for Organ Culture Samantha C.W. Chan1, Benjamin Gantenbein-Ritter1 1ARTORG Center for Biomedical Engineering, University of Bern This protocol illustrates a harvesting technique for coccygeal bovine intervertebral discs for organ culture for in vitro organ culture. Medicine Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects Camillo Sherif*1,2,3, Javier Fandino*2,4, Salome Erhardt2,4, Antonio di Ieva5, Monika Killer6, Guenther Kleinpeter1, Serge Marbacher2,4 1Department of Neurosurgery, Hospital Rudolfstiftung, 2Department of Neurosurgery, Kantonsspital Aarau, 3Cerebrovascular Research Group, Department of Biomedical Research, Medical University of Vienna, 4Cerebrovascular Research Group, Department of Intensive Care Medicine, University of Berne, 5Centre for Anatomy and Cell Biology, Department of Systematic Anatomy, Medical University of Vienna, 6Department of Neurology, Paracelsus University Salzburg An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.