Oturum Aç

McGill University

56 ARTICLES PUBLISHED IN JoVE

image

Biology

Batch Immunostaining for Large-Scale Protein Detection in the Whole Monkey Brain
Shahin Zangenehpour 1,2, Mark W. Burke 2, Avi Chaudhuri 3, Maurice Ptito 2
1Cognitive Neuroscience Unit, Montreal Neurological Institute, 2Ècole d’Optomètrie, Universitè de Montrèal, 3Department of Psychology, McGill University

Large-scale immunodetection of target proteins across the entire primate brain is possible by employing novel tissue embedding and sectioning methods combined with the use of creative apparatus for batch staining of multiple free-floating sections at a given time.

image

Biology

The Microfluidic Probe: Operation and Use for Localized Surface Processing
Cecile M. Perrault 1, Mohammad A. Qasaimeh 1, David Juncker 1
1Department of Biomedical Engineering, McGill University

In this video we present the microfluidic probe1 (MFP). We explain in detail how to assemble the MFP, mount it atop an inverted microscope, and align it relative to the substrate surface, and finally show how to use it to process a substrate surface immersed in a buffer solution.

image

Neuroscience

Dissection and Culture of Commissural Neurons from Embryonic Spinal Cord
Sébastien D. Langlois *1,2, Steves Morin *1, Patricia T. Yam 1,3,4, Frédéric Charron 1,2,5,6,7
1Molecular Biology of Neural Development, Institut de Recherches Cliniques de Montréal, 2Division of Experimental Medicine and Program in Neuroengineering, McGill University, 3Program in Neuroengineering, McGill University, 4Montreal Neurological Institute, 5Department of Anatomy and Cell Biology, McGill University, 6Department of Biology, McGill University, 7Department of Medicine, Universite de Montreal - University of Montreal

This video demonstrates a method to dissect and culture commissural neurons from E13 rat dorsal spinal cord. Dissociated commissural neurons are useful to study the cellular and molecular mechanisms of axon growth and guidance.

image

Neuroscience

Live-imaging of PKC Translocation in Sf9 Cells and in Aplysia Sensory Neurons
Carole A. Farah 1, Wayne S. Sossin 1
1Neurology and Neurosurgery, McGill University

In this video, we demonstrate visualization of PKC translocation in living cells using fluorescently tagged PKCs.

image

Biology

Collecting Variable-concentration Isothermal Titration Calorimetry Datasets in Order to Determine Binding Mechanisms
Lee A. Freiburger 1, Anthony K. Mittermaier 1, Karine Auclair 1
1Department of Chemistry, McGill University

ITC is a powerful tool for studying the binding of a ligand to its host. In complex systems however, several models may fit the data equally well. The method described here provides a means to elucidate the appropriate binding model for complex systems and extract the corresponding thermodynamic parameters.

image

Biology

Evaluation of Respiratory System Mechanics in Mice using the Forced Oscillation Technique
Toby K. McGovern *1, Annette Robichaud *2, Liah Fereydoonzad 2, Thomas F. Schuessler 2, James G. Martin 1
1Meakins-Christie Laboratories, Department of Medicine, McGill University, 2SCIREQ Scientific Respiratory Equipment Inc.

The present protocol provides a detailed step-by-step description of the procedures required to execute measurements of respiratory system mechanics as well as the assessment of airway responsiveness to inhaled methacholine in mice using the forced oscillation technique (flexiVent; SCIREQ Inc, Montreal, Qc, Canada).

image

Bioengineering

Permeabilization of Adhered Cells Using an Inert Gas Jet
Scott Cooper 1, Paul Jonak 1, Guillaume Chouinard-Pelletier 1, Sylvain Coulombe 1, Elizabeth Jones 1, Richard L. Leask 1,2
1Chemical Engineering, McGill University, 2Montreal Heart Institute

This protocol describes a method for the temporary permeabilization of adherent cells using an inert gas jet. This technique facilitates the transfer of genetic material and biomolecules into adherent mammalian cells by the utilization of mechanical forces to disrupt the plasma membrane.

image

Biology

Dithranol as a Matrix for Matrix Assisted Laser Desorption/Ionization Imaging on a Fourier Transform Ion Cyclotron Resonance Mass Spectrometer
Cuong H. Le 1,2, Jun Han 1, Christoph H. Borchers 1,2
1University of Victoria-Genome BC Proteomics Centre, University of Victoria, 2Department of Biochemistry and Microbiology, University of Victoria

Dithranol (DT; 1,8-dihydroxy-9,10-dihydroanthracen-9-one) has previously been reported as a MALDI matrix for tissue imaging of small molecules; protocols for the use of DT for the MALDI imaging of endogenous lipids on the surface of tissue sections by positive-ion MALDI-MS on an ultrahigh-resolution quadrupole-FTICR instrument are provided here.

image

Behavior

Measuring Sensitivity to Viewpoint Change with and without Stereoscopic Cues
Jason Bell 1,2, Edwin Dickinson 2, David R. Badcock 2, Frederick A. A. Kingdom 3
1Research School of Psychology, Australian National University, 2School of Psychology, University of Western Australia, 3McGill Vision Research, Department of Ophthalmology, McGill University

We discuss a novel method forviewpoint-rotation of visual stimuli, and demonstrate using a mirror stereoscopethe three-dimensional percept of rotation-in-depth. The technique can be used to investigate the role of stereoscopic cues in encoding viewpoint-rotated figures.

image

Biology

Easy Measurement of Diffusion Coefficients of EGFP-tagged Plasma Membrane Proteins Using k-Space Image Correlation Spectroscopy
Eva C. Arnspang 1, Jennifer S. Koffman 1, Saw Marlar 1, Paul W. Wiseman 2, Lene N. Nejsum 1
1Institute of Molecular Biology and Genetics and Interdisciplinary Nanoscience Center, Aarhus University, 2Departments of Chemistry and Physics, McGill University

This paper provides a step by step guide to the fluctuation analysis technique k-Space Image Correlation Spectroscopy (kICS) for measuring diffusion coefficients of fluorescently labeled plasma membrane proteins in live mammalian cells.

image

Neuroscience

Assessment of Vascular Regeneration in the CNS Using the Mouse Retina
Khalil Miloudi 1, Agnieszka Dejda 2, François Binet 3, Eric Lapalme 2, Agustin Cerani 2, Przemyslaw Sapieha 1,2,3
1Department of Neurology-Neurosurgery, McGill University, 2Department of Biochemistry, Maisonneuve-Rosemont Hospital Research Centre, University of Montréal, 3Department of Ophthalmology, Maisonneuve-Rosemont Hospital Research Centre, University of Montréal

The rodent retina has long been recognized as an accessible window to the brain. In this technical paper we provide a protocol that employs the mouse model of oxygen-induced retinopathy to study the mechanisms that lead to failure of vascular regeneration within the central nervous system after ischemic injury. The described system can also be harnessed to explore strategies to promote regrowth of functional blood vessels within the retina and CNS.

image

Environment

Colorimetric Paper-based Detection of Escherichia coli, Salmonella spp., and Listeria monocytogenes from Large Volumes of Agricultural Water
Bledar Bisha 1, Jaclyn A. Adkins 2, Jana C. Jokerst 3, Jeffrey C. Chandler 1, Alma Pérez-Méndez 4, Shannon M. Coleman 4, Adrian O. Sbodio 5, Trevor V. Suslow 5, Michelle D. Danyluk 6, Charles S. Henry 2, Lawrence D. Goodridge 7
1Department of Animal Science, University of Wyoming, 2Department of Chemistry, Colorado State University, 3Department of Environmental and Radiological Health Sciences, Colorado State University, 4Department of Animal Sciences, Colorado State University, 5Department of Plant Sciences, University of California, Davis, 6Department of Food Science and Human Nutrition, Citrus Research and Education Center, University of Florida, 7Department of Food Science and Agricultural Chemistry, McGill University

A protocol involving integrated concentration, enrichment, and end-point colorimetric detection of foodborne pathogens in large volumes of agricultural water is presented here. Water is filtered through Modified Moore Swabs (MMS), enriched with selective or non-selective media, and detection is performed using paper-based analytical devices (µPAD) imbedded with bacterial-indicative colorimetric substrates.

image

Biology

Polysome Fractionation and Analysis of Mammalian Translatomes on a Genome-wide Scale
Valentina Gandin 1, Kristina Sikström 2, Tommy Alain 3, Masahiro Morita 3, Shannon McLaughlan 1, Ola Larsson 2, Ivan Topisirovic 1
1Lady Davis Institute and Department of Oncology, McGill University, 2Department of Oncology-Pathology, Karolinska Institutet, 3Goodman Cancer Centre and Department of Biochemistry, McGill University

Ribosomes play a central role in protein synthesis. Polyribosome (polysome) fractionation by sucrose density gradient centrifugation allows direct determination of translation efficiencies of individual mRNAs on a genome-wide scale. In addition, this method can be used for biochemical analysis of ribosome- and polysome-associated factors such as chaperones and signaling molecules.

image

Neuroscience

The Use of Magnetic Resonance Spectroscopy as a Tool for the Measurement of Bi-hemispheric Transcranial Electric Stimulation Effects on Primary Motor Cortex Metabolism
Sara Tremblay 1, Vincent Beaulé 1, Sébastien Proulx 2, Louis-Philippe Lafleur 1, Julien Doyon 1, Małgorzata Marjańska 3, Hugo Théoret 1
1Department of Psychology, University of Montréal, 2Montreal Neurological Institute, McGill University, 3Center for Magnetic Resonance Research and Department of Radiology, University of Minnesota

This article aims to describe a basic protocol for combining transcranial direct current stimulation (tDCS) with proton magnetic resonance spectroscopy (1H-MRS) measurements to investigate the effects of bilateral stimulation on primary motor cortex metabolism.

image

Biology

RNA Catalyst as a Reporter for Screening Drugs against RNA Editing in Trypanosomes
Houtan Moshiri *1,2, Vaibhav Mehta *1,2, Reza Salavati 1,2,3
1Department of Biochemistry, McGill University, 2Institute of Parasitology, McGill University, 3McGill Centre for Bioinformatics, McGill University

A highly sensitive ribozyme-based assay, applicable to high-throughput screening of chemicals targeting the unique process of RNA editing in trypanosomatid pathogens, is described in this paper. Inhibitors can be used as tools for hypothesis-driven analysis of the RNA editing process and ultimately as therapeutics.

image

Neuroscience

High-resolution In Vivo Manual Segmentation Protocol for Human Hippocampal Subfields Using 3T Magnetic Resonance Imaging
Julie Winterburn 1,2, Jens C. Pruessner 3, Chavez Sofia 4,5, Mark M. Schira 6,7, Nancy J. Lobaugh 4,8, Aristotle N. Voineskos 5,9, M. Mallar Chakravarty 1,2
1Institute of Biomaterials and Biomedical Engineering, University of Toronto, 2Computational Brain Anatomy Laboratory, Douglas Institute, McGill University, 3McGill Centre for Studies in Aging, McGill University, 4MRI Unit, Research Imaging Centre, Campbell Family Mental Health Research Institute, Centre for Addiction and Mental Health, 5Department of Psychiatry, University of Toronto, 6School of Psychology, University of Wollongong, 7Neuroscience Research Australia, 8Department of Medicine, University of Toronto, 9Kimel Family Translational Imaging Genetics Research Laboratory, Research Imaging Centre, Campbell Family Mental Health Research Institute, Centre for Addiction and Mental Health

The goal of this manuscript is to study the hippocampus and hippocampal subfields using MRI. The manuscript describes a protocol for segmenting the hippocampus and five hippocampal substructures: cornu ammonis (CA) 1, CA2/CA3, CA4/dentate gyrus, strata radiatum/lacunosum/moleculare, and subiculum.

image

JoVE Core

Investigating the Effects of Antipsychotics and Schizotypy on the N400 Using Event-Related Potentials and Semantic Categorization
Vivian Gu 1, Ola Mohamed Ali 1, Katherine L'Abbée Lacas 2, J. Bruno Debruille 3,4
1Department of Psychology, McGill University, 2Department of Cognitive Science, McGill University, 3Douglas Institute, Department of Psychiatry, McGill University, 4Neurology and Neurosurgery Department, McGill University

Using event-related EEG potentials (ERPs), we investigate the effects of antipsychotic medications on abnormal semantic brain activations in healthy individuals with schizotypal traits. We use ERPs to track distinct changes in brain activity, shedding insight into the cognitive processes associated with semantic categorization.

image

Biology

Electrophoretic Mobility Shift Assay (EMSA) for the Study of RNA-Protein Interactions: The IRE/IRP Example
Carine Fillebeen 1,2, Nicole Wilkinson 1,2, Kostas Pantopoulos 1,2
1Lady Davis Institute for Medical Research, Jewish General Hospital, 2Department of Medicine, McGill University

Here we present a protocol to analyze RNA/protein interactions. The electrophoretic mobility shift assay (EMSA) is based on the differential migration of RNA/protein complexes and free RNA during native gel electrophoresis. By using a radiolabeled RNA probe, RNA/protein complexes can be visualized by autoradiography.

image

Immunology and Infection

Simplified Human Neutrophil Extracellular Traps (NETs) Isolation and Handling
Sara Najmeh *1, Jonathan Cools-Lartigue *1, Betty Giannias 1, Jonathan Spicer 1, Lorenzo E. Ferri 1
1LD MacLean Surgical Research Laboratories, Department of Surgery, McGill University

In the following protocol, we describe a very simple way to isolate Neutrophil Extracellular traps (NETs) from human whole blood using readily available reagents. We then demonstrate how the isolated NETs can be used in an in vitro adhesion assay with cancer cells.

image

Behavior

Paw-Dragging: a Novel, Sensitive Analysis of the Mouse Cylinder Test
R. Brian Roome 1,2, Jacqueline L. Vanderluit 1
1BioMedical Sciences, Faculty of Medicine, Memorial University of Newfoundland, 2Neurosciences, Faculty of Medicine, McGill University

Classical forelimb asymmetry analysis of the cylinder test is routinely used to assess behavioural deficits in rats following brain injury or stroke; however, it fails to detect consistent deficits in mice. This study demonstrates that quantifying paw-dragging behaviour is a more sensitive analysis of brain injury in mice.

image

Neuroscience

Quantitative Analysis of Climbing Defects in a Drosophila Model of Neurodegenerative Disorders
Surya T. Madabattula 1, Joel C. Strautman 1, Andrew M. Bysice 1, Julia A. O’Sullivan 1, Alaura Androschuk 1, Cory Rosenfelt 1, Kacy Doucet 1, Guy Rouleau 2, Francois Bolduc 1
1Department of Pediatrics, University of Alberta, 2Montreal Neurological Institute and Hospital, McGill University

We present an optimized inexpensive and reliable negative geotaxis assay in Drosophila melanogaster as a model for neurodegenerative disorders. Being more sensitive to mild locomotor defects, this assay will help screen for potential genetic interactions and drug targets.

image

Biology

Measuring Oxidative Stress Resistance of Caenorhabditis elegans in 96-well Microtiter Plates
Elite Possik 1,2, Arnim Pause 1,2
1Goodman Cancer Research Center, McGill University, 2Department of Biochemistry, McGill University

C. elegans is an attractive model organism to study signal transduction pathways involved in oxidative stress resistance. Here we provide a protocol to measure oxidative stress resistance of C. elegans animals in liquid phase, using several oxidizing agents in 96 well plates.

image

Neuroscience

Somatosensory Event-related Potentials from Orofacial Skin Stretch Stimulation
Takayuki Ito 1,2,3, David J. Ostry 1,4, Vincent L. Gracco 1,5
1Haskins Laboratories, 2Speech and Cognition Department, Gipsa-lab, CNRS, 3Univ. Grenoble-Alpes, 4Department of Psychology, McGill University, 5School of Communication Science and Disorders, McGill University

This paper introduces a method for obtaining somatosensory event-related potentials following orofacial skin stretch stimulation. The current method can be used to evaluate the contribution of somatosensory afferents to both speech production and speech perception.

image

Immunology and Infection

Cortical Actin Flow in T Cells Quantified by Spatio-temporal Image Correlation Spectroscopy of Structured Illumination Microscopy Data
George Ashdown 1, Elvis Pandžić 3, Andrew Cope 2, Paul Wiseman 4, Dylan Owen 1
1Department of Physics and Randall Division of Cell and Molecular Biophysics, King's College London, 2Academic Department of Rheumatology, Centre for Molecular and Cellular Biology of Inflammation, Division of Immunology, Infection and Inflammatory Disease, King's College London, 3ARC Centre for Advanced Molecular Imaging, Australian Centre for NanoMedicine, University of New South Wales Australia, 4Departments of Chemistry and Physic, McGill University

To investigate flow velocities and directionality of filamentous-actin at the T cell immunological synapse, live-cell super-resolution imaging is combined with total internal reflection fluorescence and quantified with spatio-temporal image correlation spectroscopy.

image

Behavior

An Experimental Analysis of Children's Ability to Provide a False Report about a Crime
Joshua Wyman 1, Ida Foster 1, Victoria Talwar 1
1Department of Educational and Counselling Psychology, McGill University

The current methodology is designed to provide an ecologically relevant approach for measuring the veracity, length and quality of children's true and false testimonies. Implications of the current methodology for future research and professionals who interview children will also be discussed.

image

Genetics

siRNA Transfection and EMSA Analyses on Freshly Isolated Human Villous Cytotrophoblasts
Adjimon Gatien Lokossou 1, Chirine Toufaily 2, Amandine Vargas 3, Benoit Barbeau 1
1Department of Biological Sciences, University of Quebec in Montreal, 2Department of Pharmacology and Therapeutics, McGill University, 3Department of Clinical Sciences, University of Montreal

This protocol describes a method for efficiently transfecting siRNA in freshly isolated human villous cytotrophoblasts using microporation and identifying DNA-protein complexes in these cells. Transfected cells can be monitored by Western blot and EMSA analyses during the 4-day culture time.

image

Chemistry

Highly Stable, Functional Hairy Nanoparticles and Biopolymers from Wood Fibers: Towards Sustainable Nanotechnology
Amir Sheikhi 1,2,3, Han Yang 1,2,3, Md. Nur Alam 1,2,3, Theo G. M. van de Ven 1,2,3
1Department of Chemistry, McGill University, 2Center for Self-Assembled Chemical Structures (CSACS), McGill University, 3Pulp and Paper Research Center, McGill University

Synthesis schemes to prepare highly stable wood fiber-based hairy nanoparticles and functional cellulose-based biopolymers have been detailed.

image

Biology

Preparation of Formalin-fixed Paraffin-embedded Tissue Cores for both RNA and DNA Extraction
Palak G. Patel 1,2, Shamini Selvarajah 1,2, Suzanne Boursalie 1,2, Nathan E. How 1,2, Joshua Ejdelman 3, Karl-Philippe Guerard 3, John M. Bartlett 4, Jacques Lapointe 3, Paul C. Park 1, John B. A. Okello 1,2, David M. Berman 1,2
1Department of Pathology & Molecular Medicine, Queen's University, 2Division of Cancer Biology & Genetics, Queen's Cancer Research Institute, Queen's University, 3Department of Surgery, Division of Urology, McGill University, 4Transformative Pathology Program, Ontario Institute for Cancer Research (OICR)

This modified extraction protocol improves RNA and DNA yields from more precisely targeted regions of interest in histopathologic tissue blocks.

image

Immunology and Infection

Evaluation of the Efficacy And Toxicity of RNAs Targeting HIV-1 Production for Use in Gene or Drug Therapy
Robert J. Scarborough 1,2, Kelsey L. Adams 1,2, Olivier Del Corpo 1,2, Aïcha Daher 1, Anne Gatignol 1,2,3
1Virus-Cell Interactions Laboratory, Lady Davis Institute for Medical Research, 2Department of Microbiology & Immunology, McGill University, 3Department of Medicine, McGill University

Methods to evaluate the efficacy and toxicity of RNA molecules targeting post-integration steps of the HIV-1 replication cycle are described. These methods are useful for screening new molecules and optimizing the format of existing ones.

image

Bioengineering

Mammalian Cell Encapsulation in Alginate Beads Using a Simple Stirred Vessel
Corinne A. Hoesli 1, Roger L. J. Kiang 2, Kamini Raghuram 2, René G. Pedroza 3, Karen E. Markwick 1, Antonio M. R. Colantuoni 1, James M. Piret 2
1Department of Chemical Engineering, McGill University, 2Michael Smith Laboratories & Department of Chemical and Biological Engineering, University of British Columbia, 3Michael Smith Laboratories & Department of Pharmaceutical Sciences, University of British Columbia

This video and manuscript describe an emulsion-based method to encapsulate mammalian cells in 0.5% to 10% alginate beads which can be produced in large batches using a simple stirred vessel. The encapsulated cells can be cultured in vitro or transplanted for cellular therapy applications.

image

Neuroscience

Rewiring Neuronal Circuits: A New Method for Fast Neurite Extension and Functional Neuronal Connection
Margaret H. Magdesian 1,2,3, Madeleine Anthonisen 1, G. Monserratt Lopez-Ayon 1, Xue Ying Chua 1, Matthew Rigby 1, Peter Grütter 1
1Department of Physics, McGill University, 2Department of Neurology and Neurosurgery, Montreal Neurological Institute, 3Ananda Devices

This procedure describes how to rapidly initiate, extend and connect neurites organized in microfluidic chambers using poly-D-lysine-coated beads fixed to micropipettes that guide neurite elongation.

image

Neuroscience

Tuning in the Hippocampal Theta Band In Vitro: Methodologies for Recording from the Isolated Rodent Septohippocampal Circuit
Frédéric Manseau 1, Sylvain Williams 1
1Department of Psychiatry, Douglas Mental Health University Institute, McGill University

Here, we present a protocol for recording rhythmic neuronal network theta and gamma oscillations from an isolated whole hippocampal preparation. We describe the experimental steps from extraction of the hippocampus to details of field, unitary and whole-cell patch clamp recordings as well as optogenetic pacing of the theta rhythm.

image

Neuroscience

Modeling Neuronal Death and Degeneration in Mouse Primary Cerebellar Granule Neurons
Matthew Laaper 1,2, Takrima Haque 1, Ruth S. Slack 3, Arezu Jahani-Asl 1,2,4
1Lady Davis Institute for Medical Research, Jewish General Hospital, 2Integrated Program in Neuroscience, McGill University, 3Department of Cellular and Molecular Medicine, University of Ottawa, 4Department of Oncology, Faculty of Medicine, McGill University

This protocol describes a simple method for isolating and culturing primary mouse cerebral granule neurons (CGNs) from 6-7 day old pups, efficient transduction of CGNs for loss and gain of function studies, and modelling NMDA-induced neuronal excitotoxicity, low-potassium-induced cell death, DNA-damage, and oxidative stress using the same culture model.

image

Chemistry

Peptide and Protein Quantification Using Automated Immuno-MALDI (iMALDI)
Huiyan Li 1, Robert Popp 1, Bjorn Frohlich 1, Michael X. Chen 2, Christoph H. Borchers 1,3,4,5
1University of Victoria-Genome BC Proteomics Centre, 2Jewish General Hospital, McGill University, 3Dept of Biochemistry and Microbiology, University of Victoria, 4Proteomics Centre, Segal Cancer Centre, Lady Davis Institute, Jewish General Hospital, McGill University, 5Gerald Bronfman Department of Oncology, Jewish General Hospital

A protocol for the protein quantification in complex biological fluids using automated immuno-MALDI (iMALDI) technology is presented.

image

Biochemistry

Measuring Biomolecular DSC Profiles with Thermolabile Ligands to Rapidly Characterize Folding and Binding Interactions
Robert W. Harkness V 1, Philip E. Johnson 2, Anthony K. Mittermaier 1
1Department of Chemistry, McGill University, 2Department of Chemistry, York University

We present a protocol for rapid characterization of biomolecular folding and binding interactions with thermolabile ligands using differential scanning calorimetry.

image

Neuroscience

How to Find Effects of Stimulus Processing on Event Related Brain Potentials of Close Others when Hyperscanning Partners
Amanda Tardif 1,3, Ashley Chau-Morris 1,2, Zi Yue Wang 1, Ehime Takahara 1, Tim Hadjis 1,2, Jean Debruille 1, J. Bruno Debruille 1,2,3
1Douglas Mental Health University Institute, 2Department of Psychiatry, McGill University, 3Department of Neurology and Neurosurgery, McGill University

This protocol describes key steps involved in assessing the sensitivity of the brain of one person to the stimulus processing of a close other by selecting pairs of partners, recording their electroencephalogram (EEG) simultaneously and computing their event-related brain potentials (ERPs).

image

Bioengineering

Snap Chip for Cross-reactivity-free and Spotter-free Multiplexed Sandwich Immunoassays
Huiyan Li 1,2, Sebastien Bergeron 3, Heidi Larkin 1,2,3, David Juncker 1,2
1McGill University and Génome Québec Innovation Centre, 2Biomedical Engineering Department, McGill University, 3Parallex BioAssays Inc.

We demonstrate a snap chip technology for performing cross-reactivity-free multiplexed sandwich immunoassays by simply snapping two slides. A snap apparatus is used for reliably transferring reagents from microarray-to-microarray. The snap chip can be used for any biochemical reactions requiring colocalization of different reagents without cross-contamination.

image

Immunology and Infection

Deep Vein Thrombosis Induced by Stasis in Mice Monitored by High Frequency Ultrasonography
Ryan N. Rys 1, Mark D. Blostein 1,2, Catherine A. Lemarié 1,2
1Lady Davis Institute for Medical Research, Jewish General Hospital, McGill University, 2Department of Medicine, Jewish General Hospital, McGill University

The present protocol describes the steps to obtain venous thrombosis using a stasis model. In addition, we are using a non-invasive method to measure thrombus formation and resolution over time.

image

Environment

Detection of Viruses from Bioaerosols Using Anion Exchange Resin
Joshua W. Schaeffer 1, Jeffrey C. Chandler 2, Margaret Davidson 1,3, Sheryl L. Magzamen 1, Alma Pérez-Méndez 4, Stephen J. Reynolds 1, Lawrence D. Goodridge 5, John Volckens 6, Alan B. Franklin 2, Susan A. Shriner 2, Bledar Bisha 7
1High Plains Intermountain Center for Agricultural Health and Safety, Department of Environmental and Radiological Health Sciences, Colorado State University, 2National Wildlife Research Center, Wildlife Services, Animal and Plant Health Inspection Service, United States Department of Agriculture, 3Western Sydney University, 4Leprino Foods, Inc, 5Department of Food Science and Agricultural Chemistry, McGill University, 6Department of Mechanical Engineering, Colorado State University, 7Department of Animal Science, University of Wyoming

An anion exchange resin-based method, adapted to liquid impingement-based bioaerosol sampling of viruses is demonstrated. When coupled with downstream molecular detection, the method allows for facile and sensitive detection of viruses from bioaerosols.

image

Bioengineering

High Throughput Traction Force Microscopy Using PDMS Reveals Dose-Dependent Effects of Transforming Growth Factor-β on the Epithelial-to-Mesenchymal Transition
Haruka Yoshie 1, Newsha Koushki 1, Clayton Molter 1, Peter M. Siegel 2,3, Ramaswamy Krishnan 4, Allen J. Ehrlicher 1,2
1Department of Bioengineering, McGill University, 2Goodman Cancer Research Centre, McGill University, 3Department of Medicine, McGill University, 4Department of Emergency Medicine, Beth Israel Deaconess Medical Center

We present a high throughput traction force assay fabricated with silicone rubber (PDMS). This novel assay is suitable for studying physical changes in cell contractility during various biological and biomedical processes and diseases. We demonstrate this method's utility by measuring a TGF-β dependent increase in contractility during the epithelial-to-mesenchymal transition.

image

Immunology and Infection

Processing of Bronchoalveolar Lavage Fluid and Matched Blood for Alveolar Macrophage and CD4+ T-cell Immunophenotyping and HIV Reservoir Assessment
Syim Salahuddin *1,2, Elaine Thomson *1,2,3, Oussama Méziane 1,2, Omar Farnos 2, Amélie Pagliuzza 5, Nicolas Chomont 5,6, Ron Olivenstein 1, Cecilia Costiniuk 1,3,4, Mohammad-Ali Jenabian 2,3,6
1Research Institute McGill University Health Centre, 2Department of Biological Sciences, Université de Québec à Montréal, 3Department of Microbiology & Immunology, McGill University, 4Department of Medicine, Division of Infectious Diseases, McGill University, 5Centre de Recherche du Centre Hospitalier de l'Université de Montréal, 6Département de microbiologie, infectiologie et immunologie, Université de Montréal

We describe a method for processing bronchoalveolar lavage fluid and matched peripheral blood from chronically HIV-infected individuals on antiretroviral therapy to assess pulmonary HIV reservoirs. These methods result in the acquisition of highly pure CD4 T cells and alveolar macrophages that may subsequently be used for immunophenotyping and HIV DNA/RNA quantifications by ultrasensitive polymerase chain reaction.

image

JoVE Journal

Identification of Inositol Phosphate or Phosphoinositide Interacting Proteins by Affinity Chromatography Coupled to Western Blot or Mass Spectrometry
Igor Cestari 1
1Institute of Parasitology, McGill University

This protocol focuses on the identification of proteins that bind to inositol phosphates or phosphoinositides. It uses affinity chromatography with biotinylated inositol phosphates or phosphoinositides that are immobilized via streptavidin to agarose or magnetic beads. Inositol phosphate or phosphoinositide binding proteins are identified by Western blotting or mass spectrometry.

image

JoVE Core

Experimental Methods to Study Human Postural Control
Pouya Amiri 1, Abolfazl Mohebbi 1, Robert Kearney 1
1Department of Biomedical Engineering, McGill University

This article presents an experimental/analytic framework to study human postural control. The protocol provides step-by-step procedures for performing standing experiments, measuring body kinematics and kinetics signals, and analyzing the results to provide insight into the mechanisms underlying human postural control.

image

Chemistry

Solid Phase 11C-Methylation, Purification and Formulation for the Production of PET Tracers
Thomas A. Singleton *1, Mehdi Boudjemeline *1, Robert Hopewell 1, Dean Jolly 1, Hussein Bdair 1, Alexey Kostikov 1,2
1McConnell Brain Imaging Centre, Montreal Neurological Institute, McGill University, 2Department of Neurology and Neurosurgery, McGill University

We report an efficient carbon-11 radiolabeling technique to produce clinically relevant tracers for Positron Emission Tomography (PET) using solid phase extraction cartridges. 11C-methylating agent is passed through a cartridge preloaded with precursor and successive elution with aqueous ethanol provides chemically and radiochemically pure PET tracers in high radiochemical yields.

image

Genetics

Microsatellite DNA Genotyping and Flow Cytometry Ploidy Analyses of Formalin-fixed Paraffin-embedded Hydatidiform Molar Tissues
Yassemine Khawajkie 1, Nawel Mechtouf 2, Phuong Nguyen 2, Rima Slim 1,2,3
1Department of Experimental Medicine, McGill University, 2Department of Human Genetics, McGill University, 3Department of Obstetrics and Gynecology, McGill University

Hydatidiform moles are abnormal human pregnancies with heterogeneous aetiologies that can be classified according to their morphological features and parental contribution to the molar genomes. Here, protocols of multiplex microsatellite DNA genotyping and flow cytometry of formalin-fixed paraffin-embedded molar tissues are described in detail, together with results’ interpretation and integration.

image

Immunology and Infection

Immunofluorescence Staining Using IBA1 and TMEM119 for Microglial Density, Morphology and Peripheral Myeloid Cell Infiltration Analysis in Mouse Brain
Fernando González Ibanez 1,2, Katherine Picard 1,2, Maude Bordeleau 1,3, Kaushik Sharma 1,2,4, Kanchan Bisht 1,2,4, Marie-Ève Tremblay 1,2
1Axe Neurosciences, Centre de Recherche du CHU de Québec-Université Laval, 2Département de Médecine Moléculaire, Faculté de Médecine, Université Laval, 3Integrated Program in Neuroscience, McGill University, 4Center for Brain Immunology and Glia (BIG), University of Virginia

This protocol describes a step-by-step workflow for immunofluorescent costaining of IBA1 and TMEM119, in addition to analysis of microglial density, distribution, and morphology, as well as peripheral myeloid cell infiltration in mouse brain tissue.

image

Cancer Research

Detection of Lung Tumor Progression in Mice by Ultrasound Imaging
Nour Ghaddar 1,2, Shuo Wang 1, Véronique Michaud 1, Urszula Kazimierczak 1,3, Nicolas Ah-son 1, Antonis E. Koromilas 1,4
1Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, 2Division of Experimental Medicine, Faculty of Medicine, McGill University, 3Department of Cancer Immunology, Chair of Medical Biotechnology, Poznan University of Medical Sciences, 4Gerald Bronfman Department of Oncology, Faculty of Medicine, McGill University

This protocol describes the steps taken to induce KRAS lung tumors in mice as well as the quantification of formed tumors by ultrasound imaging. Small tumors are visualized in early timepoints as B-lines. At later timepoints, relative tumor volume measurements are achieved by the measurement tool in the ultrasound software.

image

Biology

Isolating Myofibrils from Skeletal Muscle Biopsies and Determining Contractile Function with a Nano-Newton Resolution Force Transducer
Martijn van de Locht 1, Josine M. de Winter 1, Dilson E. Rassier 2, Michiel H.B. Helmes 1,3, Coen A.C. Ottenheijm 1
1Department of Physiology, Amsterdam UMC, 2Department of Kinesiology and Physical Education, Faculty of Education, McGill University, 3IONOptix BV

Presented here is a protocol to assess the contractile properties of striated muscle myofibrils with nano-Newton resolution. The protocol employs a setup with an interferometry-based, optical force probe. This setup generates data with a high signal-to-noise ratio and enables the assessment of the contractile kinetics of myofibrils.

image

Biochemistry

A Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) Platform for Investigating Peptide Biosynthetic Enzymes
Yeganeh Habibi 1, Christopher J. Thibodeaux 1
1Department of Chemistry, McGill University

Lanthipeptide synthetases catalyze multistep reactions during the biosynthesis of peptide natural products. Here, we describe a continuous, bottom-up, hydrogen-deuterium exchange mass spectrometry (HDX-MS) workflow that can be employed to study the conformational dynamics of lanthipeptide synthetases, as well as other similar enzymes involved in peptide natural product biosynthesis.

image

Neuroscience

Reliable Acquisition of Electroencephalography Data during Simultaneous Electroencephalography and Functional MRI
Hui Ming Khoo 1, Yuya Fujita 1, Naoki Tani 1, Tetsuya Shimokawa 2, Natalja Zazubovits 3, Satoru Oshino 1, Jean Gotman 3, Haruhiko Kishima 1
1Department of Neurosurgery, Osaka University Graduate School of Medicine, 2Center for Information and Neural Networks, National Institute of Information and Communications Technology, 3Montreal Neurological Institute and Hospital, McGill University

This article provides a straightforward protocol for acquiring good quality electroencephalography (EEG) data during simultaneous EEG and functional magnetic resonance imaging by utilizing readily available medical products.

image

Immunology and Infection

C-section of Preclinical Animal Model of Chorioamnionitis Triggered by Group B Streptococcus (GBS)
Taghreed A. Ayash 1, Seline Y. Vancolen 2, Marie-Julie Allard *1, Guillaume Sébire *1
1Department of Pediatrics, McGill University, 2Department of Pharmacology, McGill University

The goal of this protocol is to describe a preclinical animal model of Group B Streptococcus (GBS)-induced chorioamnionitis. The study is designed to investigate mechanistic processes, potential causal links with developmental impairments, and finally to develop translational anti-inflammatory placento- and neuro-protective treatments.

image

Medicine

Oxygenation-sensitive Cardiac MRI with Vasoactive Breathing Maneuvers for the Non-invasive Assessment of Coronary Microvascular Dysfunction
Elizabeth Hillier 1,2, Jason Covone 1, Matthias G. Friedrich 1,3
1Faculty of Medicine and Health Sciences, McGill University, 2Faulty of Medicine and Dentistry, University of Alberta, 3Departments of Medicine and Diagnostic Radiology, McGill University

The assessment of microvascular function by oxygenation-sensitive cardiac magnetic resonance imaging in combination with vasoactive breathing maneuvers is unique in its ability to assess rapid dynamic changes in myocardial oxygenation in vivo and, thus, may serve as a critically important diagnostic technique for coronary vascular function.

image

Cancer Research

Somatic Genome-Engineered Mouse Models Using In Vivo Microinjection and Electroporation
Keerthana Harwalkar 1,2, Nobuko Yamanaka 1,3, Yojiro Yamanaka 1,2,3
1Rosalind and Morris Goodman Cancer Research Institute, McGill University, 2Department of Human Genetics, McGill University, 3McGill Integrated Core for Animal Modeling (MICAM), McGill University

This protocol describes microinjection and in vivo electroporation for regionally restricted CRISPR-mediated genome editing in the mouse oviduct.

image

Neuroscience

Social Defeat Stress Model for Adolescent C57BL/6 Male and Female Mice
Andrea Harée Pantoja-Urbán 1, Samuel Richer 1, Michel Giroux 2, Dominique Nouel 2, Cecilia Flores 2,3,4,5
1Integrated Program in Neuroscience, McGill University, 2Douglas Mental Health University Institute, McGill University, 3Department of Psychiatry, McGill University, 4Department of Neurology and Neurosurgery, Faculty of Medicine, McGill University, 5Ludmer Centre for Neuroinformatics & Mental Health, McGill University

We developed an accelerated social defeat stress model for adolescent C57BL/6 mice, which works in both males and females and allows exposure during discrete adolescent periods. Exposure to this model induces social avoidance, but only in a subset of defeated male and female mice.

image

Engineering

Development of a Bio-Hybrid Mosquito Stinger-Based Atomic Force Microscopy Probe
Nicolas Juncos Ljubich 1, Justin Puma 1, Zi Xin Zhang 1, Jianyu Li 1, Changhong Cao 1
1Department of Mechanical Engineering, McGill University

Quantitative and controlled investigations into insect biting behaviors are crucial for devising effective strategies to combat vector-borne diseases. In this context, a method for fabricating a bio-hybrid atomic force microscopy (AFM) probe is introduced.

image

Behavior

Strategies for Assessing Autistic-Like Behaviors in Mice
Ziying Huang 1, Shane Wiebe 1, Laura Marsal-García 1, Ilse Gantois 1, Nahum Sonenberg 1
1Rosalind and Morris Goodman Cancer Institute, Department of Biochemistry, McGill University

Rodent models are valuable tools for studying core behaviors related to autism spectrum disorder (ASD). In this article, we expound on two behavioral tests for modeling the core features of ASD in mice: self-grooming, which assesses repetitive behavior, and the three-chamber social interaction test, which documents social impairments.

JoVE Logo

Gizlilik

Kullanım Şartları

İlkeler

Araştırma

Eğitim

JoVE Hakkında

Telif Hakkı © 2020 MyJove Corporation. Tüm hakları saklıdır