University of Technology Sydney 13 articles published in JoVE Biology Gas Chromatography-Mass Spectrometry-Based Targeted Metabolomics of Hard Coral Samples Jennifer L. Matthews1, Natasha Bartels1, Sheik Nadeem Elahee Doomun2, Simon K. Davy3, David P. De Souza2 1Climate Change Cluster (C3), University of Technology Sydney, 2Metabolomics Australia, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, 3School of Biological Sciences, Victoria University of Wellington Here, we present the extraction and preparation of polar and semi-polar metabolites from a coral holobiont, as well as separated coral host tissue and Symbiodiniaceae cell fractions, for gas chromatography-mass spectrometry analysis. Chemistry Extraction of Non-Protein Amino Acids from Cyanobacteria for Liquid Chromatography-Tandem Mass Spectrometry Analysis Sercan Pravadali-Cekic1, Jake P. Violi1, Simon M. Mitrovic2, Kenneth J. Rodgers2, David Bishop1 1Hyphenated Mass Spectrometry Laboratory (HyMaS), University of Technology Sydney, 2School of Life Sciences, University of Technology Sydney The present protocol describes the extraction of non-protein amino acids from biological matrices via trichloroacetic acid (TCA) protein precipitation and acid hydrolysis before analysis using liquid chromatography-tandem mass spectrometry. Bioengineering Cardiac Spheroids as in vitro Bioengineered Heart Tissues to Study Human Heart Pathophysiology Poonam Sharma1,2,3,4, Carmine Gentile2,3,4 1University of Newcastle, 2University of Sydney, 3Kolling Institute of Medical Research, Royal North Shore Hospital, 4University of Technology, Sydney This protocol aims to fabricate 3D cardiac spheroids (CSs) by co-culturing cells in hanging drops. Collagen-embedded CSs are treated with doxorubicin (DOX, a cardiotoxic agent) at physiological concentrations to model heart failure. In vitro testing using DOX-treated CSs may be used to identify novel therapies for heart failure patients. Biology Tethered Bilayer Lipid Membranes to Monitor Heat Transfer between Gold Nanoparticles and Lipid Membranes Amani Alghalayini1,2, Lele Jiang1, Xi Gu3, Guan Heng Yeoh3, Charles G. Cranfield1,2, Victoria Timchenko3, Bruce A. Cornell2,4, Stella M. Valenzuela1,2,5 1School of Life Sciences, University of Technology Sydney, 2ARC Research Hub for Integrated Devices for End-user Analysis at Low-levels (IDEAL), Faculty of Science, University of Technology Sydney, 3School of Mechanical and Manufacturing Engineering, The University of New South Wales, 4Surgical Diagnostics Pty Ltd., 5Institute for Biomedical Materials and Devices, University of Technology Sydney This work outlines a protocol to achieve dynamic, non-invasive monitoring of heat transfer from laser-irradiated gold nanoparticles to tBLMs. The system combines impedance spectroscopy for the real-time measurement of conductance changes across the tBLMs, with a horizontally focused laser beam that drives gold nanoparticle illumination, for heat production. Bioengineering Transplantation of a 3D Bioprinted Patch in a Murine Model of Myocardial Infarction Christopher D. Roche1,2,3,4, Carmine Gentile1,2,3 1The University of Sydney, 2University of Technology Sydney (UTS), 3The Royal North Shore Hospital, 4University Hospital of Wales This protocol aims to transplant a 3D bioprinted patch onto the epicardium of infarcted mice modeling heart failure. It includes details regarding anesthesia, the surgical chest opening, permanent ligation of the left anterior descending (LAD) coronary artery and application of a bioprinted patch onto the infarcted area of the heart. Environment In Situ Chemotaxis Assay to Examine Microbial Behavior in Aquatic Ecosystems Estelle E. Clerc1, Jean-Baptiste Raina2, Bennett S. Lambert3, Justin Seymour2, Roman Stocker1 1Institute of Environmental Engineering, Department of Civil, Environmental and Geomatic Engineering, ETH Zürich, 2Climate Change Cluster, University of Technology Sydney, 3School of Oceanography, University of Washington Presented here is the protocol for an in situ chemotaxis assay, a recently developed microfluidic device that enables studies of microbial behavior directly in the environment. Engineering Quantifying the Relative Thickness of Conductive Ferromagnetic Materials Using Detector Coil-Based Pulsed Eddy Current Sensors Nalika Ulapane1, Karthick Thiyagarajan2, David Hunt2, Jaime Valls Miro2 1Melbourne School of Engineering, University of Melbourne, 2Center for Autonomous Systems, University of Technology Sydney Here, we present a protocol to quantify the relative thickness (i.e., thickness as a percentage with respect to a reference) of conductive ferromagnetic materials using detector coil-based pulsed eddy current sensors, while overcoming the calibration requirement. Chemistry Luminescence Lifetime Imaging of O2 with a Frequency-Domain-Based Camera System Maria Moßhammer*1, Vincent V. Scholz*2, Gerhard Holst3, Michael Kühl1,4, Klaus Koren5 1Marine Biological Section, Department of Biology, University of Copenhagen, 2Center for Electromicrobiology, Aarhus University, 3PCO AG, 4Climate Change Cluster, University of Technology Sydney, 5Aarhus University Centre for Water Technology, Section for Microbiology, Department of Bioscience, Aarhus University We describe the use of a novel, frequency-domain luminescence lifetime camera for mapping 2D O2 distributions with optical sensor foils. The camera system and image analysis procedures are described along with the preparation, calibration and application of sensor foils for visualizing the O2 microenvironment in the rhizosphere of aquatic plants. Chemistry Color Spot Test As a Presumptive Tool for the Rapid Detection of Synthetic Cathinones Morgan Philp1, Ronald Shimmon1, Mark Tahtouh2, Shanlin Fu1 1Centre for Forensic Science, University of Technology Sydney (UTS), 2Australian Federal Police (AFP) Here we present a simple, inexpensive, and selective chemical spot test protocol for the detection of synthetic cathinones, a class of new psychoactive substances. The protocol is suitable for use in various areas of law enforcement that encounter illicit material. Biochemistry Optimal Preparation of Formalin Fixed Samples for Peptide Based Matrix Assisted Laser Desorption/Ionization Mass Spectrometry Imaging Workflows Matthew B. O'Rourke1,2, Matthew P. Padula2, Caine Smith3, Priscilla Youssef4, Stuart Cordwell1, Paul Witting4, Greg Sutherland3, Ben Crossett1 1Mass Spectrometry Core Facility, University of Sydney, 2Proteomics Core Facility, University of Technology Sydney, 3Neuropathology Group, Discipline of Pathology, School of Medical Sciences, University of Sydney, 4Redox Biology Group, Discipline of Pathology, School of Medical Sciences, University of Sydney This protocol describes a reproducible and reliable method for the sublimation-based preparation of formalin fixed tissue destined for imaging mass spectrometry. Medicine Imaging Metals in Brain Tissue by Laser Ablation - Inductively Coupled Plasma - Mass Spectrometry (LA-ICP-MS) Dominic J. Hare1,2, Kai Kysenius3, Bence Paul4, Beate Knauer5,6, Robert W. Hutchinson7, Ciaran O'Connor7, Fred Fryer8, Tom P. Hennessey8, Ashley I. Bush2, Peter J. Crouch3, Philip A. Doble1 1Elemental Bio-imaging Facility, University of Technology Sydney, 2Florey Institute of Neuroscience and Mental Health, The University of Melbourne, 3Department of Pathology, The University of Melbourne, 4School of Earth Sciences, The University of Melbourne, 5Research School, Ruhr University, 6Department of Physiology, Monash University, 7ESI Ltd., Bozeman, 8Agilent Technologies, Mulgrave Quantitatively mapping metals in tissue by laser ablation - inductively coupled plasma - mass spectrometry (LA-ICP-MS) is a sensitive analytical technique that can provide new insight into how metals participate in normal function and disease processes. Here, we describe a protocol for quantitatively imaging metals in thin sections of mouse neurological tissue. Biology Super-resolution Imaging of the Cytokinetic Z Ring in Live Bacteria Using Fast 3D-Structured Illumination Microscopy (f3D-SIM) Lynne Turnbull*1, Michael P. Strauss*1, Andrew T. F. Liew1, Leigh G. Monahan1, Cynthia B. Whitchurch1, Elizabeth J. Harry1 1The ithree Institute, University of Technology, Sydney Spatiotemporal information about dynamic proteins inside live cells is crucial for understanding biology. A type of super-resolution microscopy called fast 3D-structured illumination microscopy (f3D-SIM) reveals unique information about the cytokinetic Z ring in bacteria: both its bead-like appearance and the rapid dynamics of FtsZ within the ring. Biology Isolation of Small Noncoding RNAs from Human Serum Samantha Khoury1, Pamela Ajuyah1, Nham Tran2,3 1School of Medical and Molecular Biosciences, Faculty of Science, University of Technology, Sydney, 2Centre for Health Technologies, Faculty of Engineering and Information Technology, University of Technology, Sydney, 3The Sydney Head and Neck Cancer Institute, Sydney Cancer Centre, Royal Prince Alfred Hospital This protocol describes a method for extracting small RNAs from human serum. We have used this method to isolate microRNAs from cancer serum for use in DNA arrays and also singleplex quantitative PCR. The protocol utilizes phenol and guanidinium thiocyanate reagents with modifications to yield high quality RNA.
