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University of Technology Sydney

13 ARTICLES PUBLISHED IN JoVE

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Chemistry

Tuning a Parallel Segmented Flow Column and Enabling Multiplexed Detection
Sercan Pravadali-Cekic 1, Danijela Kocic 1, Stanley Hua 1, Andrew Jones 1, Gary R. Dennis 1, R. Andrew Shalliker 1
1School of Science and Health, University of Western Sydney

Here, we present a protocol for the operation and tuning of parallel segmented flow chromatography columns to enable multiplexed detection.

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Chemistry

Post Column Derivatization Using Reaction Flow High Performance Liquid Chromatography Columns
Andrew Jones 1, Sercan Pravadali-Cekic 1, Stanley Hua 1, Danijela Kocic 1, Michelle Camenzuli 2, Gary Dennis 1, Andrew Shalliker 1
1School of Science and Health, University of Western Sydney, 2Van′t Hoff Institute for Molecular Sciences, University of Amsterdam

A protocol for the use of reaction flow high performance liquid chromatography columns for methods employing post column derivatization (PCD) is presented.

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Chemistry

Curtain Flow Column: Optimization of Efficiency and Sensitivity
Sercan Pravadali-Cekic 1, Danijela Kocic 1, Stanley Hua 1, Andrew Jones 1, Gary Dennis 1, Andrew Shalliker 1
1School of Science and Health, University of Western Sydney

Here, we present a protocol for the operation and optimization of Active Flow Technology (AFT) column in Curtain flow (CF) mode for enhanced separation performance.

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Medicine

Imaging Metals in Brain Tissue by Laser Ablation - Inductively Coupled Plasma - Mass Spectrometry (LA-ICP-MS)
Dominic J. Hare 1,2, Kai Kysenius 3, Bence Paul 4, Beate Knauer 5,6, Robert W. Hutchinson 7, Ciaran O'Connor 7, Fred Fryer 8, Tom P. Hennessey 8, Ashley I. Bush 2, Peter J. Crouch 3, Philip A. Doble 1
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.

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Biochemistry

Optimal Preparation of Formalin Fixed Samples for Peptide Based Matrix Assisted Laser Desorption/Ionization Mass Spectrometry Imaging Workflows
Matthew B. O'Rourke 1,2, Matthew P. Padula 2, Caine Smith 3, Priscilla Youssef 4, Stuart Cordwell 1, Paul Witting 4, Greg Sutherland 3, Ben Crossett 1
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.

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Immunology and Infection

Development and Validation of an Ultrasensitive Single Molecule Array Digital Enzyme-linked Immunosorbent Assay for Human Interferon-α
Alba Llibre *1,2, Vincent Bondet *1,2, Mathieu P. Rodero 3, David Hunt 4, Yanick J. Crow 3,5, Darragh Duffy 1,2
1Immunobiology of Dendritic Cells, Institut Pasteur, 2INSERM U1223, 3Laboratory of Neurogenetics and Neuroinflammation, INSERM UMR1163, Institut Imagine, 4MRC Human Genetics Unit, MRC Institute of Genetics and Molecular Medicine, University of Edinburgh, 5Manchester Centre for Genomic Medicine, University of Manchester

Here we present a protocol to describe the development and validation of a single molecule array digital ELISA assay, which enables the ultra-sensitive detection of all IFN-α subtypes in human samples.

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JoVE Core

Quantifying the Relative Thickness of Conductive Ferromagnetic Materials Using Detector Coil-Based Pulsed Eddy Current Sensors
Nalika Ulapane 1, Karthick Thiyagarajan 2, David Hunt 2, Jaime Valls Miro 2
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.

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Chemistry

Luminescence Lifetime Imaging of O2 with a Frequency-Domain-Based Camera System
Maria Moßhammer *1, Vincent V. Scholz *2, Gerhard Holst 3, Michael Kühl 1,4, Klaus Koren 5
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.

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Environment

In Situ Chemotaxis Assay to Examine Microbial Behavior in Aquatic Ecosystems
Estelle E. Clerc 1, Jean-Baptiste Raina 2, Bennett S. Lambert 3, Justin Seymour 2, Roman Stocker 1
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.

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Biology

Tethered Bilayer Lipid Membranes to Monitor Heat Transfer between Gold Nanoparticles and Lipid Membranes
Amani Alghalayini 1,2, Lele Jiang 1, Xi Gu 3, Guan Heng Yeoh 3, Charles G. Cranfield 1,2, Victoria Timchenko 3, Bruce A. Cornell 2,4, Stella M. Valenzuela 1,2,5
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.

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Biology

Temporary Translocation of Entire Mistletoe Plants to Understand the Mechanistic Basis of Animal Foraging Decisions
Melinda Cook 1, Andy Leigh 1, David M. Watson 2
1School of Life Sciences, University of Technology Sydney, 2Gulbali Institute, School of Agricultural, Environmental, and Veterinary Science, Charles Sturt University

This work outlines a simple experimental procedure to quantify behavioral drivers of foraging decisions in free-living animals, temporarily relocating mistletoe plants to novel locations and measuring visitation rates.

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Chemistry

Extraction of Non-Protein Amino Acids from Cyanobacteria for Liquid Chromatography-Tandem Mass Spectrometry Analysis
Sercan Pravadali-Cekic 1, Jake P. Violi 1, Simon M. Mitrovic 2, Kenneth J. Rodgers 2, David Bishop 1
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.

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Biology

Gas Chromatography-Mass Spectrometry-Based Targeted Metabolomics of Hard Coral Samples
Jennifer L. Matthews 1, Natasha Bartels 1, Sheik Nadeem Elahee Doomun 2, Simon K. Davy 3, David P. De Souza 2
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.

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