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Keele University

6 ARTICLES PUBLISHED IN JoVE

Neuroscience

Flow Cytometry Protocols for Surface and Intracellular Antigen Analyses of Neural Cell Types

Vishal Menon¹, Ria Thomas^1,2, Arun R. Ghale^1,3, Christina Reinhard¹, Jan Pruszak^1,4

¹Emmy Noether-Group for Stem Cell Biology, Department of Molecular Embryology, Institute of Anatomy and Cell Biology, University of Freiburg, ²Spemann Graduate School of Biology and Medicine and Faculty of Biology, University of Freiburg, ³School of Life Sciences, Keele University, ⁴Center for Biological Signaling Studies (BIOSS), University of Freiburg

We provide a detailed description of a protocol for flow cytometric analysis of surface antigens and/or intracellular antigens in neural cell types. Critical aspects of experimental planning, step-by-step methodological procedures, and fundamental principles of flow cytometry are explained in order to enable neurobiologists to exploit this powerful technology.

Intra-articular injection of murine MSC for the treatment of inflammation and tissue degradation in Antigen-induced Arthritis.

Alasdair Kay¹, Jim Middleton², Oksana Kehoe³

¹Institute for Science and Technology in Medicine, Keele University, ²Faculty of Medicine and Dentistry, University of Bristol, ³Keele University

Here, we present a protocol to induce inflammatory arthritis in vivo using antigen response, termed Antigen-induced Arthritis (AIA), an effective tool for research into Rheumatoid Arthritis (RA) research. Additionally we demonstrate the therapeutic effects of infusion of MSC to reduce inflammation, cartilage degradation and expression of TNFα.

Biochemistry

Online Size-exclusion and Ion-exchange Chromatography on a SAXS Beamline

Martha E. Brennich¹, Adam R. Round^2,3, Stephanie Hutin⁴

¹Structural Biology Group, European Synchrotron Radiation Facility, ²European Molecular Biology Laboratory, ³School of Chemical and Physical Sciences, Keele University, ⁴Groupe de Microscopie Electronique et Méthodes, Institut de Biologie Structurale

The determination of the solution structure of a protein by small angle X-ray scattering (SAXS) requires monodisperse samples. Here, we present two possibilities to ensure minimal delays between sample preparation and data acquisition: online size-exclusion chromatography (SEC) and online ion-exchange chromatography (IEC).

Medicine

Corneal Epithelial Abrasion with Ocular Burr As a Model for Cornea Wound Healing

Solja Kalha¹, Alison Kuony¹, Frederic Michon^1,2

¹Helsinki Institute of Life Science, Institute of Biotechnology, University of Helsinki, ²School of Medicine and Institute for Science and Technology in Medicine, Keele University

This protocol describes a method to inflict an abrasion to the ocular surface of the mouse, and to follow the wound healing process thereafter. The protocol takes advantage of an ocular burr to partially remove the surface epithelium of the eye in anaesthetized mice.

Medicine

Use of a Percutaneous Ventricular Assist Device/Left Atrium to Femoral Artery Bypass System for Cardiogenic Shock

Swethika Sundaravel¹, Poonam Velagapudi¹, Mamas Mamas², Sandeep Nathan³, Alexander Truesdell⁴

¹University of Nebraska Medical Center, ²Keele Cardiovascular Research Group, Keele University, ³University of Chicago Medicine, ⁴Virginia Heart/Inova Heart and Vascular Institute

The following article describes the stepwise procedure for placement of a device (e.g., Tandemheart) in cardiogenic shock (CS) that is a percutaneous left ventricular assist device (pLVAD) and a left atrial to femoral artery bypass (LAFAB) system that bypasses and supports the left ventricle (LV) in CS.

JoVE Core

Multiplexed Fluorescent Immunohistochemical Staining of Four Endometrial Immune Cell Types in Recurrent Miscarriage

Yiwei Zhao¹, Gene Chi Wai Man^1,2, Loucia Kit Ying Chan¹, Xi Guo¹, Yingyu Liu³, Tao Zhang¹, Joseph Kwong^1,4, Chi Chiu Wang^1,5, Xiaoyan Chen³, Tin Chiu Li¹

¹Department of Obstetrics and Gynaecology, The Chinese University of Hong Kong, Prince of Wales Hospital, ²Department of Orthopaedics and Traumatology, The Chinese University of Hong Kong, Prince of Wales Hospital, ³Department of Obstetrics and Gynaecology, Shenzhen Baoan Women’s and Children’s Hospital, Shenzhen University, ⁴School of Medicine, Faculty of Medicine and Health Sciences, Keele University, ⁵Li Ka Shing Institute of Health Sciences; School of Biomedical Sciences, The Chinese University of Hong Kong

Despite the advancements in multiplex immunohistochemistry and multispectral imaging, characterizing the density and clustering of major immune cells simultaneously in the endometrium remains a challenge. This paper describes a detailed multiplex staining protocol and imaging for the simultaneous localization of four immune cell types in the endometrium.