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Dalhousie University

12 ARTICLES PUBLISHED IN JoVE

Biology

Horizontal Slice Preparation of the Retina

Ryosuke Enoki¹, Tatjana C. Jakobs², Amane Koizumi²

¹Dpt of Physiology and Biophysics, Dalhousie University, ²Massachusetts General Hospital, Harvard Medical School

Traditionally the vertical slice and the whole-mount preparation of the retina have been used to study the function of retinal circuits. Here, we describe the novel slicing method to preserve the dendritic morphology of retinal neurons intact.

Engineering

Measurement of Coherence Decay in GaMnAs Using Femtosecond Four-wave Mixing

Daniel Webber¹, Tristan de Boer¹, Murat Yildirim¹, Sam March¹, Reuble Mathew¹, Angela Gamouras¹, Xinyu Liu², Margaret Dobrowolska², Jacek Furdyna², Kimberley Hall¹

¹Department of Physics and Atmospheric Science, Dalhousie University, ²Department of Physics, University of Notre Dame

The technique of femtosecond four-wave mixing is described, including spectrally-resolved and time-resolved configurations. We illustrate the utility of this technique for the investigation of crucial physical properties in the III-V diluted magnetic semiconductors, afforded by its nonlinearity and high temporal resolution.

Neuroscience

Immunohistochemical and Calcium Imaging Methods in Wholemount Rat Retina

Allison Sargoy¹, Steven Barnes^1,2,3, Nicholas C. Brecha^1,2,4, Luis Pérez De Sevilla Müller¹

¹Department of Neurobiology, University of California, Los Angeles, ²Veterans Administration Greater Los Angeles Healthcare System, ³Departments of Physiology & Biophysics and Ophthalmology & Visual Sciences, Dalhousie University, ⁴Departments of Neurobiology and Medicine, Jules Stein Eye Institute, CURE-Digestive Diseases Research Center, David Geffen School of Medicine, University of California, Los Angeles

Immunohistochemistry protocols are used to study the localization of a specific protein in the retina. Calcium imaging techniques are employed to study calcium dynamics in retinal ganglion cells and their axons.

Neuroscience

Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis

Maximillian DJ Fiander^*1, Matthew AJ Chedrawe^*1, Anna-Claire Lamport¹, Turgay Akay², George S Robertson^1,3

¹Pharmacology, Dalhousie University, ²Medical Neuroscience, Dalhousie University, ³Psychiatry, Dalhousie University

Kinematic gait analysis in the sagittal plane yields highly precise information about how movement is executed. We describe the application of these techniques to identify gait deficits for mice subjected to autoimmune-mediated demyelination. These methods may also be used to characterize gait deficits for other mouse models featuring impaired locomotion.

Immunology and Infection

Use of Viral Entry Assays and Molecular Docking Analysis for the Identification of Antiviral Candidates against Coxsackievirus A16

Jonathan Y. Wang¹, Chien-Ju Lin², Ching-Hsuan Liu^3,4, Liang-Tzung Lin^3,5

¹Department of Molecular Biosciences, University of Texas at Austin, ²School of Pharmacy, College of Pharmacy, Kaohsiung Medical University, ³Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, ⁴Department of Microbiology and Immunology, Dalhousie University, ⁵Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University

The goal of the protocol is to illustrate the different assays relating to viral entry that can be used to identify candidate viral entry inhibitors.

Neuroscience

A Standardized Pipeline for Examining Human Cerebellar Grey Matter Morphometry using Structural Magnetic Resonance Imaging

Rebecca Kerestes¹, Shuo Han², Srinivas Balachander³, Carlos Hernandez-Castillo⁴, Jerry L. Prince^5,6, Jörn Diedrichsen⁷, Ian H. Harding^1,8

¹Department of Neuroscience, Central Clinical School, Monash University, ²Department of Biomedical Engineering, The Johns Hopkins University, ³Department of Psychiatry, National Institute of Mental Health & Neuro Sciences (NIMHANS), ⁴Faculty of Computer Science, Dalhousie University, ⁵Department of Electrical and Computer Engineering, The Johns Hopkins University, ⁶Department of Computer Science, The Johns Hopkins University, ⁷Brain and Mind Institute, Department for Statistical and Actuarial Sciences, Department for Computer Science, Western University, ⁸Monash Biomedical Imaging, Monash University

A standardized pipeline is presented for examining cerebellum grey matter morphometry. The pipeline combines high-resolution, state-of-the-art approaches for optimized and automated cerebellum parcellation and voxel-based registration of the cerebellum for volumetric quantification.

JoVE Core

Organic Solvent-Based Protein Precipitation for Robust Proteome Purification Ahead of Mass Spectrometry

Jessica L. Nickerson¹, Venus Baghalabadi¹, Ziheng Dang¹, Victoria A. Miller², Sara L. Little², Alan A. Doucette¹

¹Department of Chemistry, Dalhousie University, ²Proteoform Scientific Inc.

The present protocol describes solvent-based protein precipitation under controlled conditions for robust and rapid recovery and purification of proteome samples prior to mass spectrometry.

Immunology and Infection

Intravital Widefield Fluorescence Microscopy of Pulmonary Microcirculation in Experimental Acute Lung Injury Using a Vacuum-Stabilized Imaging System

Stefan Hall¹, Sufyan Faridi², Irene Euodia^2,3, Sophie Tanner⁴, Andrew Krzysztof Chojnacki⁵, Kamala D. Patel^6,7, Juan Zhou^2,4, Christian Lehmann^1,2,4,8

¹Department of Physiology and Biophysics, Dalhousie University, ²Department of Microbiology and Immunology, Dalhousie University, ³Department of Neuroscience, Dalhousie University, ⁴Department of Anesthesia, Pain Management, and Perioperative Medicine, Dalhousie University, ⁵Live Cell Imaging Center, University of Calgary, ⁶Department of Physiology and Pharmacology, University of Calgary, ⁷Department of Biochemistry and Molecular Biology, University of Calgary, ⁸Department of Pharmacology, Dalhousie University

Intravital fluorescence microscopy can be utilized to study leukocyte-endothelial interactions and capillary perfusion in real-time. This protocol describes methods to image and quantify these parameters in the pulmonary microcirculation using a vacuum-stabilized lung imaging system.

Environment

Enhanced Reproducibility and Precision of High-Throughput Quantification of Bacterial Growth Data Using a Microplate Reader

Leili Abkar¹, Florentin M. Wilfart², Marta Piercey³, Graham A. Gagnon⁴

¹Chemical and Biological Engineering Department, University of British Columbia, ²Process Engineering and Applied Science, Dalhousie University, ³Food Science and Technology, Dalhousie University, ⁴Center for Water Resources Studies, Civil and Resource Engineering, Dalhousie University

Here, a high-throughput protocol is presented to measure growth data, including growth curves, growth rate, and maximum growth rate. The protocol was verified and validated using two biofilm-producing bacteria. The results and approach applied in this study can be expanded to other high-throughput protocols using microplate readers.

Medicine

Large-Animal Model of Donation after Circulatory Death and Normothermic Regional Perfusion for Cardiac Assessment

Khalil Khalil^*1,2, Roberto V. P. Ribeiro^*3, Julgans S. Alvarez^4,5, Mitesh V. Badiwala^4,5, Shant Der Sarkissian^1,2, Nicolas Noiseux^1,2

¹Centre de Recherche du Centre Hospitalier de l'Université de Montréal (CRCHUM), ²Department of Surgery, Faculty of Medicine, Université de Montréal, ³Division of Cardiac Surgery, Dalhousie University, ⁴Division of Cardiovascular Surgery, Peter Munk Cardiac Centre, Toronto General Hospital, ⁵Department of Surgery, University of Toronto

The protocol describes a large-animal (porcine) model of donation after circulatory death, followed by thoracoabdominal normothermic regional perfusion that closely simulates the clinical scenario in heart transplantation, and has the potential to facilitate therapeutic studies and strategies.

Tips And Tricks for Proteome Sample Preparation

Alan A. Doucette¹

¹Department of Chemistry, Dalhousie University

Tips And Tricks for Proteome Sample Preparation

Neuroscience

Recording Forelimb Muscle Activity in Head-Fixed Mice with Chronically Implanted EMG Electrodes

Amy Claire Kristl¹, Turgay Akay², Andrew Miri¹

¹Department of Neurobiology, Northwestern University, ²Department of Medical Neuroscience, Dalhousie University

This protocol describes the hand fabrication and surgical implantation of electromyographic (EMG) electrodes in the forelimb muscles of mice to record muscle activity during head-fixed behavior experiments.