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21 ARTICLES PUBLISHED IN JoVE

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Medicine

A Simple Guide Screw Method for Intracranial Xenograft Studies in Mice
Jacqueline F. Donoghue 1, Oliver Bogler 2, Terrance G. Johns 1
1Monash Institute of Medical Research , 2MD Anderson Cancer Centre, University of Texas

In order to evaluate novel therapeutic paradigms for the treatment of glioma, physiological relevant models are essential. We utilize an implantable guide screw procedure for establishment of intracranial xenograft models that is more rapid and safer than stereotactic approaches.

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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.

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Developmental Biology

Stencil Micropatterning of Human Pluripotent Stem Cells for Probing Spatial Organization of Differentiation Fates
Geetika Sahni *1, Jun Yuan *1, Yi-Chin Toh 1,2
1Department of Biomedical Engineering, National University of Singapore, 2Singapore Institute of Neurotechnology (SINAPSE), National University of Singapore

Human pluripotent stem cells (hPSCs) have the intrinsic ability to differentiate and self-organize into distinct tissue patterns; although this requires the presentation of spatial environmental gradients. We present stencil micropatterning as a simple and robust method to generate biochemical and mechanical gradients for controlling hPSC differentiation patterns.

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Biology

Mucin Agarose Gel Electrophoresis: Western Blotting for High-molecular-weight Glycoproteins
Kathryn A. Ramsey 1,2, Zachary L. Rushton 1, Camille Ehre 1,3
1Marsico Lung Institute/CF Center, University of North Carolina at Chapel Hill, 2Telethon Kids Institute, University of Western Australia, 3Department of Pediatrics, University of North Carolina at Chapel Hill

Mucins are high-molecular-weight glycoconjugates, with size ranging from 0.2 to 200 megadalton (MDa). As a result of their size, mucins do not penetrate conventional polyacrylamide gels and require larger pores for separation. We provide a detailed protocol for mucin agarose gel electrophoresis to assess relative quantification and study polymer assembly.

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Biology

The Murine Choline-Deficient, Ethionine-Supplemented (CDE) Diet Model of Chronic Liver Injury
Jully Gogoi-Tiwari 1, Julia Köhn-Gaone 1, Corey Giles 2, Dirk Schmidt-Arras 3, Francis D. Gratte 1,4, Caryn L. Elsegood 1, Geoffrey W. McCaughan 5,6,7, Grant A. Ramm 8,9, John K. Olynyk 10,11, Janina E.E. Tirnitz-Parker 1,12
1School of Biomedical Sciences & Curtin Health Innovation Research Institute, Curtin University, 2School of Public Health & Curtin Health Innovation Research Institute, Curtin University, 3Institute of Biochemistry, Christian-Albrechts-University, 4School of Veterinary and Life Sciences, Murdoch University, 5Centenary Institute of Cancer Medicine and Cell Biology, The University of Sydney, 6Royal Prince Alfred Hospital, 7A.W. Morrow Gastroenterology and Liver Centre, 8QIMR Berghofer Medical Research Institute, 9Faculty of Medicine and Biomedical Sciences, The University of Queensland, 10Fiona Stanley and Fremantle Hospitals, 11School of Medical and Health Sciences, Edith Cowan University, 12School of Medicine and Pharmacology, University of Western Australia

Here we describe a common method to induce chronic liver injury in mice by feeding of a choline-deficient and ethionine-supplemented (CDE) diet. We demonstrate health monitoring, liver perfusion, isolation, and preservation. A time course of six weeks can inform about liver injury, pathohistology, fibrosis, inflammatory, and liver progenitor cell responses.

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Chemistry

Measurements of Long-range Electronic Correlations During Femtosecond Diffraction Experiments Performed on Nanocrystals of Buckminsterfullerene
Rebecca A. Ryan 1, Sophie Williams 1, Andrew V. Martin 1, Ruben A. Dilanian 1, Connie Darmanin 2, Corey T. Putkunz 1, David Wood 3, Victor A. Streltsov 4, Michael W.M. Jones 5, Naylyn Gaffney 6, Felix Hofmann 7, Garth J. Williams 8, Sebastien Boutet 9, Marc Messerschmidt 10, M. Marvin Seibert 11, Evan K. Curwood 11, Eugeniu Balaur 2, Andrew G. Peele 5, Keith A. Nugent 2, Harry M. Quiney 1, Brian Abbey 2
1ARC Centre of Excellence in Advanced Molecular Imaging, School of Physics, University of Melbourne, 2Australian Research Council (ARC) Centre of Excellence in Advanced Molecular Imaging, Department of Chemistry and Physics, La Trobe Institute for Molecular Sciences, La Trobe University, 3Department of Physics, Imperial College London, 4Florey Institute of Neuroscience and Mental Health, 5Science and Engineering Faculty, Queensland University of Technology, 6Swinburne University of Technology, 7Department of Engineering Science, University of Oxford, 8Brookhaven National Laboratory, 9Linac Coherent Light Source, SLAC National Accelerator Laboratory, 10BioXFEL Science and Technology Center, 11Laboratory of Molecular Biophysics, Department of Cell and Molecular Biology, Uppsala University, 12Australian Synchrotron

We describe an experiment designed to probe the electronic damage induced in nanocrystals of Buckminsterfullerene (C60) by intense, femtosecond pulses of X-rays. The experiment found that, surprisingly, rather than being stochastic, the X-ray induced electron dynamics in C60 are highly correlated, extending over hundreds of unit cells within the crystals1.

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JoVE Journal

Isolation and Respiratory Measurements of Mitochondria from Arabidopsis thaliana
Wenhui Lyu 1, Jennifer Selinski 1, Lu Li 1, David A. Day 2, Monika W. Murcha 3, James Whelan 1, Yan Wang 1
1ARC Centre of Excellence in Plant Energy Biology, Department of Animal, Plant and Soil Science, School of Life Science, La Trobe University, 2School of Biological Sciences, Flinders University, 3ARC Centre of Excellence in Plant Energy Biology, University of Western Australia

As mitochondria are only a small percentage of the plant cell, they need to be purified for a range of studies. Mitochondria can be isolated from a variety of plant organs by homogenization, followed by differential and density gradient centrifugation to obtain a highly purified mitochondrial fraction.

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Genetics

A Fast Silver Staining Protocol Enabling Simple and Efficient Detection of SSR Markers using a Non-denaturing Polyacrylamide Gel
Ling Huang *1, Xiaohui Deng *1, Ronghua Li 1, Yanshi Xia 1, Guihua Bai 2, Kadambot H.M. Siddique 3, Peiguo Guo 1
1College of Life Sciences, Guangzhou University, 2Hard Winter Wheat Genetics Research Unit, United States Department of Agriculture - Agricultural Research Service, 3The UWA Institute of Agriculture, University of Western Australia

Here, we report a simple and low-cost silver staining protocol which requires only three reagents and 7 min of processing, and is suitable for fast generation of high-quality SSR data in the genetic analysis.

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Genetics

Candidate Gene Testing in Clinical Cohort Studies with Multiplexed Genotyping and Mass Spectrometry
Sarah E. Ashley 1,2, Braydon A. Meyer 2,3, Justine A. Ellis 2,3,4, David J. Martino 2,3,5
1Molecular Genetics of Chronic Inflammation and Allergic Disease, Max-Delbrück Center for Molecular Medicine, 2Murdoch Childrens Research Institute, 3Department of Paediatrics, University of Melbourne, 4Centre for Social and Early Emotional Development, Faculty of Health, Deakin University, 5Department of Paediatrics, University of Western Australia

Identification of genetic variants contributing to complex human disease allows us to identify novel mechanisms. Here, we demonstrate a multiplex genotyping approach to candidate genes or gene pathway analysis that maximizes the coverage at low cost and is amenable to cohort-based studies.

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Medicine

Preparation, Procedures and Evaluation of Platelet-Rich Plasma Injection in the Treatment of Knee Osteoarthritis
Ziming Chen *1,2, Zhantao Deng *1, Yuanchen Ma 1, Junxing Liao 1, Qingtian Li 1, Mengyuan Li 1, Hua Liu 1, Ganghong Chen 1, Chaoming Zeng 1, Qiujian Zheng 1
1Department of Orthopedics, Guangdong General Hospital (Guangdong Academy of Medical Sciences), 2Shantou University Medical College

Knee osteoarthritis is frequently seen in the orthopedic department. We introduce in detail the entire knee osteoarthritis treatment process with platelet-rich plasma injection, including preparation, procedures, and evaluation.

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Biology

Quantification of Proliferative and Dead Cells in Enteroids
Hua-Shan Li *1, Shao-Fang Xu *1, Jian-Ying Sheng *1, Zhi-Hui Jiang 1, Jing Wang 1, Ning Ding 1, Tao Wang 1, Matthew A. Odenwald 2, Jerrold R. Turner 2,3, Wei-Qi He 1, Hong Xu 1, Juan-Min Zha 1
1Jiangsu Key Laboratory of Neuropsychiatric Diseases and Cambridge-Suda (CAM-SU) Genomic Resource Center, Medical College of Soochow University, Department of Oncology, The First Affiliated Hospital of Soochow University, 2Department of Pathology, University of Chicago, 3Department of Pathology, Brigham and Women's Hospital–Harvard Medical School

The presented protocol uses flow cytometry to quantify the number of proliferating and dead cells in cultured mouse enteroids. This method is helpful to evaluate the effects of drug treatment on organoid proliferation and survival.

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Bioengineering

Growth of Human and Sheep Corneal Endothelial Cell Layers on Biomaterial Membranes
Jennifer Walshe 1, Najla Al Khaled Abdulsalam 2, Shuko Suzuki 1, Traian V. Chirila 1,3,4,5,6,7,8, Damien G. Harkin 1,3,4
1Queensland Eye Institute, 2King Faisal University, 3School of Biomedical Sciences, Faculty of Health, Queensland University of Technology, 4Institute of Health and Biomedical Innovation, Queensland University of Technology, 5Science and Engineering Faculty, Queensland University of Technology, 6Faculty of Medicine, University of Queensland, 7Australian Institute for Bioengineering and Nanotechnology, University of Queensland, 8Faculty of Science, University of Western Australia

This protocol describes the critical steps required to establish and grow corneal endothelial cell cultures from explants of human or sheep tissue. A method for subculturing corneal endothelial cells on membranous biomaterials is also presented.

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Behavior

Implementation of a Real-Time Psychosis Risk Detection and Alerting System Based on Electronic Health Records using CogStack
Tao Wang 1, Dominic Oliver 2, Yamiko Msosa 1, Craig Colling 3, Giulia Spada 2, Łukasz Roguski 4, Amos Folarin 1, Robert Stewart 3,5, Angus Roberts 1,3, Richard J. B. Dobson 1,3,4,6, Paolo Fusar-Poli 2,3,7,8
1Department of Biostatistics and Health Informatics, Institute of Psychiatry, Psychology and Neuroscience, King's College London, 2Early Psychosis: Interventions and Clinical-detection (EPIC) lab, Department of Psychosis Studies, Institute of Psychiatry, Psychology & Neuroscience, King's College London, 3National Institute for Health Research, Maudsley Biomedical Research Centre, South London and Maudsley National Health Service (NHS) Foundation Trust, 4Institute of Health Informatics, University College London, 5Department of Psychological Medicine, Institute of Psychiatry, Psychology and Neuroscience, King's College London, 6Health Data Research UK London, University College London, 7OASIS service, South London and Maudsley National Health Service (NHS) Foundation Trust, 8Department of Brain and Behavioral Sciences, University of Pavia

We demonstrate how to deploy a real-time psychosis risk calculation and alerting system based on CogStack, an information retrieval and extraction platform for electronic health records.

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Medicine

A Mouse Model of Incompletely Resected Soft Tissue Sarcoma for Testing (Neo)adjuvant Therapies
Francois X. Rwandamuriye 1,2, Breana J. Weston 2,3, Terrance G. Johns 2, W. Joost Lesterhuis 1,2, Rachael M. Zemek 2
1School of Biomedical Sciences, University of Western Australia, 2Telethon Kids Institute, University of Western Australia, 3College of Science, Health, Engineering and Education, Murdoch University

In this protocol, we describe a mouse model of incomplete surgical resection of soft tissue sarcoma for testing (neo)adjuvant therapies.

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Medicine

Murine Precision-Cut Liver Slices as an Ex Vivo Model of Liver Biology
Michael A. Pearen *1, Hong Kiat Lim *1, Francis D. Gratte 2,3, Manuel A. Fernandez-Rojo 1,4,5, Sujeevi K. Nawaratna 6, Geoffrey N. Gobert 7, John K. Olynyk 8,9, Janina E. E. Tirnitz-Parker 2,10, Grant A. Ramm 1,4
1Hepatic Fibrosis Group, QIMR Berghofer Medical Research Institute, 2School of Pharmacy and Biomedical Sciences, Curtin Health Innovation Research Institute, Curtin University, 3School of Veterinary and Life Sciences, Murdoch University, 4School of Medicine, The University of Queensland, 5Madrid Institute for Advanced Studies (IMDEA) in Food, CEI UAM+CSIC, 6School of Medicine, Griffith University, 7School of Biological Sciences, Queen's University Belfast, 8Department of Gastroenterology & Hepatology, Fiona Stanley and Fremantle Hospital Group, 9School of Medical and Health Sciences, Edith Cowan University, 10Centre for Cell Therapy and Regenerative Medicine, School of Biomedical Sciences, University of Western Australia

This protocol provides a simple and reliable method for the production of viable precision-cut liver slices from mice. The ex vivo tissue samples can be maintained under laboratory tissue culture conditions for multiple days, providing a flexible model to examine liver pathobiology.

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Medicine

Applying a Three-dimensional Uniaxial Mechanical Stimulation Bioreactor System to Induce Tenogenic Differentiation of Tendon-Derived Stem Cells
Ziming Chen *1, Peilin Chen *1, Rui Ruan *1, Lianzhi Chen 1, Jun Yuan 1, David Wood 1, Tao Wang 1, Ming Hao Zheng 1
1Centre of Orthopaedic Translational Research, Medical School, University of Western Australia

A three-dimensional uniaxial mechanical stimulation bioreactor system is an ideal bioreactor for tenogenic-specific differentiation of tendon-derived stem cells and neo-tendon formation.

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Behavior

Controlled Rotation of Human Observers in a Virtual Reality Environment
Michael Falconbridge 1, Phillip Falconbridge 2, David R. Badcock 1
1School of Psychology, University of Western Australia, 2Falco Enterprise

The controlled physical rotation of a human observer is desirable for certain experimental, recreational, and educational applications. This paper outlines a method for converting an office swivel chair into a medium for controlled physical rotation in a virtual reality environment.

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Medicine

Continuous Telemetric In Utero Tracheal Pressure Measurements in Fetal Lambs
Mar Janna Dahl 1, Stefano Robbiani 2, Chiara Veneroni 2, Anna Lavizzari 3, Sebastian Amos 1, Gabrielle C. Musk 1,4, Helen Kershaw 4, Jonathan W. Davis 5,6, Fabio Mosca 3, Peter B. Noble 1, Raffaele Dellacà *2, J. Jane Pillow *1,6
1School of Human Sciences, The University of Western Australia, 2Dipartimento di Elettronica, Informazione e Bioingegneria, Politecnico di Milano University, 3NICU, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, 4Animal Care Services, The University of Western Australia, 5Medical School, University of Western Australia, 6Telethon Kids Institute

Optimization of occlusion pressure and duration required to improve lung hypoplasia with tracheal occlusion is essential to improve in utero corrective treatments for congenital diaphragmatic hernia (CDH). This study reports a new method for continuous measurement of the tracheal pressure in an occluded and unoccluded fetal lamb surgical model of CDH.

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Bioengineering

Decellularized Apple-Derived Scaffolds for Bone Tissue Engineering In Vitro and In Vivo
Maxime Leblanc Latour 1, Maryam Tarar 2, Ryan J. Hickey 1, Charles M. Cuerrier 1, Isabelle Catelas 3,4,5, Andrew E. Pelling 1,2,6,7
1Department of Physics, University of Ottawa, 2Department of Biology, University of Ottawa, 3Department of Mechanical Engineering, University of Ottawa, 4Department of Surgery, University of Ottawa, 5Department of Biochemistry, Microbiology and Immunology, University of Ottawa, 6Institute for Science, Society and Policy, University of Ottawa, 7SymbioticA, School of Human Sciences, University of Western Australia

In this study, we detail methods of decellularization, physical characterization, imaging, and in vivo implantation of plant-based biomaterials, as well as methods for cell seeding and differentiation in the scaffolds. The described methods allow the evaluation of plant-based biomaterials for bone tissue engineering applications.

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Chemistry

An Automated Radiosynthesis of [68Ga]Ga-FAPI-46 for Routine Clinical Use
Laurence Morandeau 1, Joseph A. Ioppolo 1,2, Eva Alvarez de Eulate 1, Shifaza Mohamed 1, Danica Cullen 1, Ali H. Asad 1, Roslyn J. Francis 2,3,4, Janette Atkinson 1
1Radiopharmaceutical Production and Development Centre (RAPID), Medical Technology and Physics Department, Sir Charles Gairdner Hospital, 2National Imaging Facility, Centre for Microscopy, Characterisation and Analysis, University of Western Australia, 3Department of Nuclear Medicine, Sir Charles Gairdner Hospital, 4Medical School, University of Western Australia

This work describes the automated production of up to 1.7 GBq of [68Ga]Ga-FAPI-46 on the iPHASE MultiSyn synthesizer for PET imaging of fibroblast activation protein.

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Medicine

Amplitude-Integrated EEG in Infants at Risk of Hypoxic-Ischemic Encephalopathy: A Feasibility Study in Road and Air Transport in Western Australia
Alexander Wilson 1,2,3, Varuna Chaudhary 1,2,3, Ela Chakkrapani 4,5, Mary O'Dea 6,7, Jonathan Davis 1,2,3,8,9,10
1Government of Western Australia, Child and Adolescent Health Service, 2Newborn Emergency Transport Service Western Australia (NETS WA), 3Perth Children's Hospital, 4University of Bristol, 5St Michael's Hospital, 6Coombe Hospital, 7Children's Health Ireland, 8Telethon Kids Institute, 9King Edward Memorial Hospital, 10University of Western Australia

Here, we present a feasibility study to assess a portable amplitude-integrated electroencephalogram (aEEG) recording system during the transport of infants with suspected hypoxic ischemic encephalopathy (HIE).

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