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University of Southampton

13 ARTICLES PUBLISHED IN JoVE

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Neuroscience

Preparation of Neuronal Co-cultures with Single Cell Precision
Ngoc-Duy Dinh 1, Ya-Yu Chiang 1,2, Heike Hardelauf 1, Sarah Waide 1, Dirk Janasek 1, Jonathan West 1,3
1Leibniz-Institut für Analytische Wissenschaften, ISAS, 2Department of Biochemical Engineering, University College London, 3Institute for Life Sciences, University of Southampton

Protocols for single neuron microfluidic arraying and water masking for the in-chip plasma patterning of biomaterial coatings are described. Highly interconnected co-cultures can be prepared using minimal cell inputs.

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Chemistry

The Use of Gas Chromatography to Analyze Compositional Changes of Fatty Acids in Rat Liver Tissue during Pregnancy
Helena L. Fisk 1, Annette L. West 1, Caroline E. Childs 1, Graham C. Burdge 1, Philip C. Calder 1
1Human Development & Health Academic Unit, Faculty of Medicine, University of Southampton

Pregnancy leads to significant changes to the fatty acid composition of maternal tissues. Lipid profiles can be obtained via gas chromatography to allow identification and quantification of fatty acids in individual lipid classes among rats fed various high and low fat diets during pregnancy.

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Medicine

Measurement of Dynamic Scapular Kinematics Using an Acromion Marker Cluster to Minimize Skin Movement Artifact
Martin B. Warner 1, Paul H. Chappell 2, Maria J. Stokes 1
1Faculty of Health Sciences, University of Southampton, 2Electronics and Computer Sciences, University of Southampton

This report presents details of how to adopt the acromion marker cluster method of obtaining scapular kinematics when using a passive marker motion capture device. As has been described in the literature, this method provides a robust, non-invasive, three-dimensional, dynamic and valid measurement of scapular kinematics, minimizing skin movement artifact.

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Developmental Biology

Derivation of Cardiac Progenitor Cells from Embryonic Stem Cells
Ieng Lam Lei 1, Lei Bu 2, Zhong Wang 1
1Cardiac Surgery, University of Michigan, 2Leon H Charney Division of Cardiology, New York University School of Medicine

In this protocol, derivation of cardiac progenitor cells from both mouse and human embryonic stem cells will be illustrated. A major strategy in this protocol is to enrich cardiac progenitor cells with flow cytometry using fluorescent reporters engineered into the embryonic stem cell lines.

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Medicine

Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome
Alicja M. Baczynska 1,2, Sarah Shaw 3, Helen C. Roberts 1,2,3,5, Cyrus Cooper 2,3,4, Avan Aihie Sayer 1,2,3,5,6, Harnish P. Patel 1,2,3
1Academic Geriatric Medicine, University of Southampton, University Hospital Southampton, 2National Institute for Health Research Southampton Biomedical Research Center, University of Southampton and University Hospital Southampton NHS Foundation Trust, 3MRC Lifecourse Epidemiology Unit, University of Southampton, 4National Institute for Health Research Musculoskeletal Biomedical Research Unit, University of Oxford, 5National Institute for Health Research Collaboration for Leadership in Applied Health Research and Care, 6Newcastle University Institute of Ageing and Institute of Health and Society, Newcastle University

This video demonstrates the technique of percutaneous muscle biopsy of the human vastus lateralis using the Weil-Blakesley conchotome.

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Engineering

Resonance Raman Spectroscopy of Extreme Nanowires and Other 1D Systems
David C. Smith 1, Joseph H. Spencer 1, Jeremy Sloan 2, Liam P. McDonnell 1, Harrison Trewhitt 2, Reza J. Kashtiban 2, Eric Faulques 3
1School of Physics and Astronomy, University of Southampton, 2Department of Physics, University of Warwick, 3Institut des Matériaux Jean Rouxel, University of Nantes CNRS

The paper describes a method for producing extreme nanowires by melt infiltration into carbon nanotubes and how 1D systems may be characterized and investigated using Resonance Raman Spectroscopy to determine vibrational and optical excitation energies.

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Neuroscience

Dorsal Root Ganglion Injection and Dorsal Root Crush Injury as a Model for Sensory Axon Regeneration
Menghon Cheah 1, James W. Fawcett 1, Melissa R. Andrews 2,3
1John van Geest Center for Brain Repair, University of Cambridge, 2School of Medicine, University of St. Andrews, 3Department of Biological Sciences, University of Southampton

This protocol presents the use of a dorsal root ganglion (DRG) injection with a viral vector and a concurrent dorsal root crush injury in an adult rat as a model to study sensory axon regeneration. This model is suitable for investigating the use of gene therapy to promote sensory axon regeneration.

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Medicine

Surgical Treatment for Benign Prostatic Hyperplasia: Holmium Laser Enucleation of the Prostate (HoLEP).
Huan Xu *1, Xiang Wan *1, Meng Gu 1, Yanbo Chen 1, Qiling Shi 1, Qi Chen 1, Zhong Wang 1
1Department of Urology, Shanghai 9th People's Hospital, Shanghai Jiaotong University School of Medicine

Here we present a safe and efficient protocol, holmium laser enucleation of the prostate, to treat benign prostatic hyperplasia.

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Cancer Research

A GPC3-targeting Bispecific Antibody, GPC3-S-Fab, with Potent Cytotoxicity
Yanlan Wang 1,2, Jiayu Liu 1,2, Haitao Pan 1,2, Jieyu Xing 1,2, Xiaoqiong Wu 1,2, Qing Li 1,2, Zhong Wang 1,2
1School of Pharmaceutical Sciences, Sun Yat-Sen University, 2Center for Cellular & Structural Biology, Sun Yat-Sen University

Here, we present a protocol to produce a bispecific antibody GPC3-S-Fab in Escherichia coli. The purified GPC3-S-Fab has potent cytotoxicity against GPC3 positive liver cancer cells.

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Chemistry

A New Straightforward Method for Lipophilicity (logP) Measurement using 19F NMR Spectroscopy
Zhong Wang 1, Benjamin F. Jeffries 1, Hannah R. Felstead 1, Neil J. Wells 1, Elisabetta Chiarparin 2, Bruno Linclau 1
1Chemistry, University of Southampton, 2Medicinal Chemistry, Oncology, IMED Biotech Unit, AstraZeneca

A novel and straightforward variation of the shake-flask method was developed for accurate lipophilicity measurement of fluorinated compounds by 19F NMR spectroscopy.

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Biochemistry

Fixed Target Serial Data Collection at Diamond Light Source
Sam Horrell 1, Danny Axford 1, Nicholas E. Devenish 1, Ali Ebrahim 1, Michael A. Hough 2, Darren A. Sherrell 1,3, Selina L. S. Storm 1,4, Ivo Tews 5, Jonathan A. R. Worrall 2, Robin L. Owen 1
1Diamond Light Source, Harwell Science and Innovation Campus, 2School of Life Sciences, University of Essex, 3X-ray Science Division, Argonne National Laboratory, 4European Molecular Biology Laboratory, Hamburg Outstation c/o DESY, 5Biological Sciences, Institute for Life Sciences, University of Southampton

We present a comprehensive guide to fixed target sample preparation, data collection, and data processing for serial synchrotron crystallography at Diamond beamline I24.

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Medicine

In vitro Assessment of Cardiac Reprogramming by Measuring Cardiac Specific Calcium Flux with a GCaMP3 Reporter
Zhaokai Li *1,2, Liu Liu *1, Zhong Wang 1
1Department of Cardiac Surgery, Cardiovascular Center, The University of Michigan, Ann Arbor, 2Department of Cardiovascular Medicine, Xiangya Hospital, Central South University

We describe here, the establishment and application of an Tg(Myh6-cre)1Jmk/J /Gt(ROSA)26Sortm38(CAG-GCaMP3)Hze/J (referred to as αMHC-Cre/Rosa26A-Flox-Stop-Flox-GCaMP3 below) mouse reporter line for cardiac reprogramming assessment. Neonatal cardiac fibroblasts (NCFs) isolated from the mouse strain are converted into induced cardiomyocytes (iCMs), allowing for convenient and efficient evaluation of reprogramming efficiency and functional maturation of iCMs via calcium (Ca2+) flux.

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Bioengineering

Optical Photothermal Infrared - Fluorescence In Situ Hybridization (OPTIR-FISH)
Zhongyue Guo *1, Yeran Bai *2,3, Fátima C. Pereira 4, Ji-Xin Cheng 1,5
1Department of Biomedical Engineering, Boston University, 2Neuroscience Research Institute, Department of Molecular, Cellular, and Developmental Biology, University of California Santa Barbara, 3Photothermal Spectroscopy Corp., 4School of Biological Sciences, University of Southampton, 5Department of Electrical & Computer Engineering, Photonics Center, Boston University

Here, we present a protocol using optical photothermal infrared-fluorescence in situ hybridization (OPTIR-FISH), also known as mid-infrared photothermal-FISH (MIP-FISH), to identify individual cells and understand their metabolism. This methodology can be applied broadly for diverse applications, including mapping cellular metabolism with single-cell resolution.

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