University of Auckland View Institution's Website 15 articles published in JoVE Bioengineering Simultaneous Brightfield, Fluorescence, and Optical Coherence Tomographic Imaging of Contracting Cardiac Trabeculae Ex Vivo Jarrah M. Dowrick1, Alex J. Anderson1, Ming L. Cheuk1, Kenneth Tran1, Poul M. F. Nielsen1,2, June-Chiew Han1, Andrew J. Taberner1,2 1Auckland Bioengineering Institute, University of Auckland, 2Department of Engineering Science, University of Auckland This protocol presents a collection of sarcomere, calcium, and macroscopic geometric data from an actively contracting cardiac trabecula ex vivo. These simultaneous measurements are made possible by the integration of three imaging modalities. Chemistry Electrochemical Preparation of Poly(3,4-Ethylenedioxythiophene) Layers on Gold Microelectrodes for Uric Acid-Sensing Applications Mahsa - Motshakeri1, Anthony R. J. Phillips2,3, Paul A. Kilmartin1 1School of Chemical Sciences, The University of Auckland, 2School of Biological Sciences, The University of Auckland, 3Surgical and Translational Research Center, The University of Auckland We describe aqueous and organic solvent systems for the electropolymerization of poly(3,4-ethylenedioxythiophene) to create thin layers on the surface of gold microelectrodes, which are used for sensing low molecular weight analytes. Biology Small-Scale Plasma Membrane Preparation for the Analysis of Candida albicans Cdr1-mGFPHis Golnoush Madani*1, Erwin Lamping*1, Hee Ji Lee1, Masakazu Niimi1,2, Alok K. Mitra3, Richard D. Cannon1 1Sir John Walsh Research Institute, Faculty of Dentistry, University of Otago, 2Department of Microbiology, Faculty of Medicine, Chulalongkorn University, 3School of Biological Sciences, University of Auckland This article presents a small-scale plasma membrane isolation protocol for the characterization of Candida albicans ABC (ATP-binding cassette) protein Cdr1, overexpressed in Saccharomyces cerevisiae. A protease-cleavable C-terminal mGFPHis double tag with a 16-residue linker between Cdr1 and the tag was designed to facilitate the purification and detergent-screening of Cdr1. Medicine Characterization of a Novel Human Organotypic Retinal Culture Technique Charisse Y. J. Kuo*1, Henry H. Louie1, Ilva D. Rupenthal1, Odunayo O. Mugisho*1 1Buchanan Ocular Therapeutics Unit, Department of Ophthalmology and the New Zealand National Eye Centre, University of Auckland This study aims to develop a novel human organotypic retinal culture (HORC) model that prevents compromising retinal integrity during explant handling. This is achieved by culturing the retina with the overlying vitreous and the underlying retinal pigment epithelium-choroid (RPE-choroid) and sclera. Developmental Biology A Simplified Method for Generating Kidney Organoids from Human Pluripotent Stem Cells Aneta Przepiorski1, Amanda E. Crunk1, Teresa M. Holm2, Veronika Sander2, Alan J. Davidson2, Neil A. Hukriede1,3 1Department of Developmental Biology, University of Pittsburgh, School of Medicine, 2Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, 3Center for Critical Care Nephrology, University of Pittsburgh, School of Medicine Here we describe a protocol to generate kidney organoids from human pluripotent stem cells (hPSCs). This protocol generates kidney organoids within two weeks. The resulting kidney organoids can be cultured in large-scale spinner flasks or multi-well magnetic stir plates for parallel drug-testing approaches. Engineering High Throughput Analysis of Liquid Droplet Impacts Matheu A.J. Broom1, Geoff R. Willmott1,2 1The Department of Physics and The MacDiarmid Institute for Advanced Materials and Nanotechnology, The University of Auckland, 2School of Chemical Sciences, The University of Auckland This protocol enables efficient collection of experimental high-speed images of liquid drop impacts, and speedy analysis of those data in batches. To streamline these processes, the method describes how to calibrate and set up apparatus, generate an appropriate data structure, and deploy an image analysis script. Neuroscience Determining the Functional Status of the Corticospinal Tract Within One Week of Stroke Marie-Claire Smith1,2,3, Suzanne J. Ackerley1,2, Emma J. Monigatti3, Benjamin J. Scrivener3, Cathy M. Stinear1,2 1Department of Medicine, University of Auckland, 2Centre for Brain Research, University of Auckland, 3Allied Health, Auckland District Health Board This protocol is for evaluating corticospinal tract function within 1 week of stroke. It can be used to select and stratify patients in trials of interventions designed to improve upper limb motor recovery and outcomes and in clinical practice for predicting upper limb functional outcomes 3 months after stroke. Immunology and Infection Targeting Drugs to Larval Zebrafish Macrophages by Injecting Drug-Loaded Liposomes Tanja Linnerz1, Manju Kanamala2, Jonathan W. Astin1, Nicola Dalbeth3, Zimei Wu2, Christopher J. Hall1 1Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, University of Auckland, 2School of Pharmacy, Faculty of Medical and Health Sciences, University of Auckland, 3School of Medicine, Faculty of Medical and Health Sciences, University of Auckland Here, we describe the synthesis of drug-loaded liposomes and their microinjection into larval zebrafish for the purpose of targeting drug delivery to macrophage-lineage cells. Cancer Research In Vivo Imaging and Quantitation of the Host Angiogenic Response in Zebrafish Tumor Xenografts Denver D. Britto1, Christopher J. Hall1, Jonathan W. Astin1 1Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland The aim of this method is to generate an in vivo model of tumor angiogenesis by xenografting mammalian tumor cells into a zebrafish embryo that has fluorescently-labelled blood vessels. By imaging the xenograft and associated vessels, a quantitative measurement of the angiogenic response can be obtained. Behavior Classical Short-Delay Eyeblink Conditioning in One-Year-Old Children Lucy K. Goodman1, Nicola S. Anstice1,2, Suzanne Stevens3, Benjamin Thompson1,4, Trecia A. Wouldes3 1School of Optometry and Vision Science, The University of Auckland, 2Discipline of Optometry, University of Canberra, 3Department of Psychological Medicine, The University of Auckland, 4School of Optometry and Vision Science, University of Waterloo This protocol describes an eyeblink conditioning paradigm appropriate for experiments with one-year-old infants. Commercial or custom-made equipment can be used to deliver the stimuli, and data collection and analysis should be performed on the video recordings. Bioengineering Creating a Structurally Realistic Finite Element Geometric Model of a Cardiomyocyte to Study the Role of Cellular Architecture in Cardiomyocyte Systems Biology Vijay Rajagopal1,2,3, Gregory Bass2,3, Shouryadipta Ghosh1,2,3, Hilary Hunt2,4, Cameron Walker5, Eric Hanssen6, Edmund Crampin2,3,4,7,8, Christian Soeller9 1Cell Structure and Mechanobiology Group, University of Melbourne, 2Systems Biology Laboratory, Melbourne School of Engineering, University of Melbourne, 3Department of Biomedical Engineering, University of Melbourne, 4School of Mathematics and Statistics, Faculty of Science, University of Melbourne, 5Department of Engineering Science, University of Auckland, 6Advanced Microscopy Facility, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, 7ARC Centre of Excellence in Convergent Bio-Nano Science and Technology, University of Melbourne, 8School of Medicine, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, 9Living Systems Institute, University of Exeter This protocol outlines a novel method to create a spatially detailed finite element model of the intracellular architecture of cardiomyocytes from electron microscopy and confocal microscopy images. The power of this spatially detailed model is demonstrated using case studies in calcium signaling and bioenergetics. Biology Visualization and Quantification of Mesenchymal Cell Adipogenic Differentiation Potential with a Lineage Specific Marker Jennifer Eom1, Vaughan Feisst1, Louis Ranjard2, Kerry Loomes1, Tanvi Damani1, Victoria Jackson-Patel1,3, Michelle Locke4,5, Hilary Sheppard1, Pritika Narayan1,6, P. Rod Dunbar1,7 1School of Biological Sciences, The University of Auckland, 2Research School of Biology, The Australian National University, 3Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, The University of Auckland, 4Department of Plastic Surgery, Counties Manukau District Health Board, 5Department of Surgery, Faculty of Medical and Health Sciences, The University of Auckland, 6Biomedical Imaging Research Unit, Faculty of Medical and Health Sciences, The University of Auckland, 7Maurice Wilkins Centre, The University of Auckland Traditional methods of assessing adipogenic differentiation are cheap and easy to use, but are not specific to changes in gene expression. We have developed an assay to quantify mesenchymal cell differentiation into mature adipocytes using a lineage specific marker. This assay has diverse applications across basic research and clinical medicine. Neuroscience Recording Brain Electromagnetic Activity During the Administration of the Gaseous Anesthetic Agents Xenon and Nitrous Oxide in Healthy Volunteers Andria Pelentritou1, Levin Kuhlmann1, John Cormack2, Will Woods3, Jamie Sleigh4, David Liley1 1Centre for Human Psychopharmacology, Swinburne University of Technology, 2 Simultaneous magnetoencephalography and electroencephalography provides a useful tool to search for common and distinct macro-scale mechanisms of reductions in consciousness induced by different anesthetics. This paper illustrates the empirical methods underlying the recording of such data from healthy humans during N-Methyl-D-Aspartate-(NMDA)-receptor-antagonist-based anesthesia during inhalation of nitrous oxide and xenon. Neuroscience Paired Whole Cell Recordings in Organotypic Hippocampal Slices Chantelle Fourie1, Marianna Kiraly2, Daniel V. Madison*2, Johanna M. Montgomery*1 1Department of Physiology and Centre for Brain Research, University of Auckland, 2Department of Molecular and Cellular Physiology, Stanford University Pair recordings are simultaneous whole cell patch clamp recordings from two synaptically connected neurons, enabling precise electrophysiological and pharmacological characterization of the synapses between individual neurons. Here we describe the detailed methodology and requirements for establishing this technique in organotypic hippocampal slice cultures in any laboratory equipped for electrophysiology. Medicine The Measurement and Treatment of Suppression in Amblyopia Joanna M. Black1, Robert F. Hess2, Jeremy R. Cooperstock3, Long To3, Benjamin Thompson1 1Department of Optometry and Vision Science, University of Auckland, 2Department of Ophthalmology, McGill University, 3Centre for Intelligent Machines, McGill University Amblyopia is a developmental disorder of the visual cortex that is often accompanied by strong suppression of one eye. We present a new technique for measuring and treating interocular suppression in patients with amblyopia that can be deployed using virtual reality goggles or a portable iPod Touch device.
