University of Missouri, Columbia View Institution's Website 24 articles published in JoVE Bioengineering Troubleshooting and Quality Assurance in Hyperpolarized Xenon Magnetic Resonance Imaging: Tools for High-Quality Image Acquisition Ummul Afia Shammi1, Gabriela María Garcίa Delgado2, Robert Thomen2,3 1Roy Blunt NextGen Precision Health Center, University of Missouri, 2Chemical and Biomedical Engineering, University of Missouri, 3Radiology, School of Medicine, University of Missouri Here, we present a protocol for obtaining high-quality hyperpolarized xenon-129 magnetic resonance images, covering hardware, software, data acquisition, sequence selection, data management, k-space utilization, and noise analysis. Biology Improved Methods for Preparing Transverse Sections and Unrolled Whole Mounts of Maize Leaf Primordia for Fluorescence and Confocal Imaging Janlo M. Robil1,2, Cedric David T. Cortez1, Chloe Margalaux R. Villafuerte1, Edgar Daniel C. Dela Peña1, Derreck O. De Leon1, Roselle Joy R. Rioja1, Paula McSteen2 1Department of Biology, School of Science and Engineering, Ateneo de Manila University, 2Division of Biological Sciences, Interdisciplinary Plant Group, and Missouri Maize Center, University of Missouri Maize leaf primordia are deeply ensheathed and rolled, making them difficult to study. Here, we present methods for preparing transverse sections and unrolled whole mounts of maize leaf primordia for fluorescence and confocal imaging. Developmental Biology Multi-Photon Laser Ablation of Cytoplasmic Microtubule Organizing Centers in Mouse Oocytes Daniela Londoño-Vásquez1,2, Alexander Jurkevich3, Ahmed Z. Balboula1 1Animal Sciences Research Center, University of Missouri, 2Universidad Nacional de Colombia, 3Advanced Light Microscopy Core, University of Missouri An optimized protocol is presented that enables the depletion of cytoplasmic microtubule organizing centers in mouse oocytes during metaphase I using a near-infrared femtosecond laser. Neuroscience Imaging Mitochondrial Ca2+ Uptake in Astrocytes and Neurons using Genetically Encoded Ca2+ Indicators (GECIs) Nannan Zhang1, Zhe Zhang1,2, Ilker Ozden2, Shinghua Ding1,2 1Dalton Cardiovascular Research Center, University of Missouri-Columbia, 2Department of Biomedical, Biological and Chemical Engineering, University of Missouri-Columbia This proctocol aims to provide a method for in vitro and in vivo mitochondrial Ca2+ imaging in astrocytes and neurons. Immunology and Infection Measurement of Pulse Propagation Velocity, Distensibility and Strain in an Abdominal Aortic Aneurysm Mouse Model Neekun Sharma*1,3, Zhe Sun*2,3, Michael A. Hill1,2,3, Chetan P. Hans1,2,3 1Division of Cardiovascular Medicine, University of Missouri, 2Medical Pharmacology and Physiology, University of Missouri, 3Dalton Cardiovascular Research Center, University of Missouri This manuscript describes a detailed protocol for using high frequency ultrasound imaging to measure luminal diameter, pulse propagation velocity, distensibility and radial strain on a mouse model of abdominal aortic aneurysm. Environment Using Deuterium Oxide as a Non-Invasive, Non-Lethal Tool for Assessing Body Composition and Water Consumption in Mammals Sarah E. Hooper1, Amanda N. Eshelman2, Ashley N. Cowan3, Alicia Roistacher4, Tyler S. Paneitz2, Sybill K. Amelon5 1Department of Veterinary Pathobiology, University of Missouri, 2College of Veterinary Medicine, University of Missouri, 3Department of Animal Science, University of Missouri, 4School of Natural Resources, University of Missouri, 5USDA USFS Northern Research Station This article describes the deuterium oxide dilution technique in two mammals, an insectivore and carnivore, to determine total body water, lean body mass, body fat mass, and water consumption. Cancer Research An Oncogenic Hepatocyte-Induced Orthotopic Mouse Model of Hepatocellular Cancer Arising in the Setting of Hepatic Inflammation and Fibrosis Xiaoqiang Qi*1,2,3, Emily Schepers*1,2, Diego Avella1,2, Eric T. Kimchi1,2, Jussuf T. Kaifi1,2, Kevin F. Staveley-O'Carroll1,2, Guangfu Li1,2,3 1Department of Surgery, University of Missouri-Columbia, 2Ellis Fischel Cancer Center, University of Missouri-Columbia, 3Molecular Microbiology and Immunology, University of Missouri-Columbia Here, we describe the development of a clinically relevant murine model of liver cancer recapitulating the typical immune features of hepatocellular cancer (HCC). Biology Live Cell Fluorescence Microscopy to Observe Essential Processes During Microbial Cell Growth Matthew Howell1, Jeremy J. Daniel1, Pamela J.B. Brown1 1Division of Biological Sciences, University of Missouri Understanding the function of essential processes in bacteria is challenging. Fluorescence microscopy with target-specific dyes can provide key insights into microbial cell growth and cell cycle progression. Here, Agrobacterium tumefaciens is used as a model bacterium to highlight methods for live cell imaging for characterization of essential processes. Behavior Assessing Working Memory in Children: The Comprehensive Assessment Battery for Children – Working Memory (CABC-WM) Kathryn Cabbage1, Shara Brinkley2, Shelley Gray2, Mary Alt3, Nelson Cowan4, Samuel Green5, Trudy Kuo6, Tiffany P. Hogan1 1Communication Sciences and Disorders, MGH Institute of Health Professions, 2Speech and Hearing Science, Arizona State University, 3Speech, Language, and Hearing Sciences, University of Arizona, 4Department of Psychological Sciences, University of Missouri-Columbia, 5Sanford School of Social and Family Dynamics, Arizona State University, 6School of Social and Behavioral Sciences, New College of Interdisciplinary Arts and Sciences, Arizona State University - West Working memory predicts a significant amount of variance for a variety of cognitive tasks, including speaking, reading, and writing. However, few tools are available to assess working memory in children. We present an innovative, computer-based battery that comprehensively assesses different components of working memory in school-age children. Engineering Evanescent Field Based Photoacoustics: Optical Property Evaluation at Surfaces Benjamin S. Goldschmidt1, Anna M. Rudy2, Charissa A. Nowak3, Yowting Tsay3, Paul J. D. Whiteside3, Heather K. Hunt3 1Biomedical Engineering, Duquesne University, 2Department of Computer Science, University of Missouri, 3Department of Bioengineering, University of Missouri Here we present a protocol to estimate material and surface optical properties using the photoacoustic effect combined with total internal reflection. This technique evanescent field-based photoacoustics can be used to create a photoacoustic metrology system to estimate materials' thicknesses, bulk and thin film refractive indices, and explore their optical properties. Biology Hydroponics: A Versatile System to Study Nutrient Allocation and Plant Responses to Nutrient Availability and Exposure to Toxic Elements Nga T. Nguyen1, Samuel A. McInturf1, David G. Mendoza-Cózatl1 1Division of Plant Sciences, C.S. Bond Life Sciences Center, University of Missouri, Columbia Here, we present an easy-to-follow protocol to establish a successful hydroponic system for plant nutrition studies. This protocol has been extensively tested in Arabidopsis and can easily be adapted to other plant species to study specific nutritional requirements or the effect of non-essential elements on plant growth and development. Chemistry Facile and Efficient Preparation of Tri-component Fluorescent Glycopolymers via RAFT-controlled Polymerization Wei Wang1, John M. Lester1, Anthony E. Amorosa2, Deborah L. Chance3, Valeri V. Mossine1, Thomas P. Mawhinney1,4 1Department of Biochemistry, University of Missouri, 2Department of Biological Sciences, University of Missouri, 3Department of Molecular Microbiology & Immunology, University of Missouri, 4Department of Child Health, University of Missouri An efficient, three-step synthesis of RAFT-based fluorescent glycopolymers, consisting of glycomonomer preparation, copolymerization, and post-modification, is demonstrated. This protocol can be used to prepare RAFT-based statistical glycopolymers with desired structures. Biology Isolation of Microvascular Endothelial Tubes from Mouse Resistance Arteries Matthew J. Socha1, Steven S. Segal1,2 1Medical Pharmacology and Physiology, University of Missouri, 2Dalton Cardiovascular Research Center We present a preparation for visualizing and manipulating calcium signaling in native, intact microvascular endothelium. Endothelial tubes freshly isolated from mouse resistance arteries supplying skeletal muscle retain in vivo morphology and dynamic signaling within and between neighboring cells. Endothelial tubes can be prepared from microvessels of other tissues and organs. Immunology and Infection Isolation of Precursor B-cell Subsets from Umbilical Cord Blood Md Almamun1, Jennifer L. Schnabel1, Susan T. Gater2, Jie Ning1, Kristen H. Taylor1 1Department of Pathology and Anatomical Sciences, University of Missouri-Columbia, 2Laboratory for Infectious Disease Research, University of Missouri-Columbia Here we describe a protocol for isolating subsets of precursor B-cells from umbilical cord blood. A sufficient quantity and quality of nucleic acids may be extracted from the cells and used in subsequent assays utilizing DNA or RNA. Biology Evaluation of Muscle Function of the Extensor Digitorum Longus Muscle Ex vivo and Tibialis Anterior Muscle In situ in Mice Chady H. Hakim1, Nalinda B. Wasala1, Dongsheng Duan1 1Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri Changes in limb muscle contractile and passive mechanical properties are important biomarkers for muscle diseases. This manuscript describes physiological assays to measure these properties in the murine extensor digitorum longus and tibialis anterior muscles. Biology Method for the Isolation and Identification of mRNAs, microRNAs and Protein Components of Ribonucleoprotein Complexes from Cell Extracts using RIP-Chip Garrett M. Dahm1,2, Matthew M. Gubin1,2, Joseph D. Magee2, Patsharaporn Techasintana1,2, Robert Calaluce2, Ulus Atasoy1,2,3 1Molecular Microbiology and Immunology, University of Missouri, 2Department of Surgery, University of Missouri, 3Child Health, University of Missouri A step by step protocol to isolating and identifying RNA associated complexes through RIP-Chip. Bioengineering Attaching Biological Probes to Silica Optical Biosensors Using Silane Coupling Agents Carol E. Soteropulos1, Heather K. Hunt1 1Department of Biological Engineering, University of Missouri Biosensors interface with complex, biological environments and perform targeted detection by combining highly sensitive sensors with highly specific probes attached to the sensor via surface modification. Here, we demonstrate the surface functionalization of silica optical sensors with biotin using silane coupling agents to bridge the sensor and the biological environment. Biology Isolation of Primary Mouse Trophoblast Cells and Trophoblast Invasion Assay Kathleen A. Pennington1, Jessica M. Schlitt1, Laura C. Schulz1 1Obstetrics, Gynecology and Women's Health, University of Missouri In this protocol, we describe the dissection of placentae from the mouse on pregnancy d10.5, followed by isolation of trophoblast cells using a Percoll gradient. We then demonstrate use of the isolated cells in a matrigel invasion assay. Medicine Mouse Model of Surgically-induced Endometriosis by Auto-transplantation of Uterine Tissue Katherine E. Pelch1, Kathy L. Sharpe-Timms2, Susan C. Nagel1 1Obstetrics, Gynecology and Women’s Health and Division of Biological Sciences, University of Missouri, 2Obstetrics, Gynecology and Women’s Health and Animal Sciences, University of Missouri A description of the surgical induction of endometriosis in mice and rats by auto-transplantation of uterine tissue to the arterial cascade of the intestinal mesentery. Bioengineering Detection and Isolation of Circulating Melanoma Cells using Photoacoustic Flowmetry Christine M. O'Brien1, Kyle Rood1, Shramik Sengupta1, Sagar K. Gupta1, Thiago DeSouza1, Aaron Cook1, John A. Viator1 1Biological Engineering, Christopher S. Bond Life Sciences Center, Dermatology, University of Missouri We have developed a flow cytometer using laser induced ultrasound to detect circulating melanoma cells as an early indicator of metastatic disease. Medicine Delivery of Therapeutic Agents Through Intracerebroventricular (ICV) and Intravenous (IV) Injection in Mice Jacqueline J. Glascock1, Erkan Y. Osman1, Tristan H. Coady2, Ferrill F. Rose1, Monir Shababi3, Christian L. Lorson3 1Department of Molecular Microbiology and Immunology, Bond Life Sciences Center, University of Missouri, 2Department of Biological Sciences, Columbia University, 3Department of Veterinary Pathobiology, Bond Life Sciences Center, University of Missouri This article demonstrates two very different methods of injection: 1) into the brain (intracerebroventricular) and 2) systemic (intravenous) to introduce the therapeutic agents into the central nervous system of neonatal mice. Biology Microiontophoresis and Micromanipulation for Intravital Fluorescence Imaging of the Microcirculation Pooneh Bagher1, Luis Polo-Parada1,2, Steven S. Segal1,2 1Department of Medical Pharmacology and Physiology, University of Missouri, 2Dalton Cardiovascular Research Center, University of Missouri Microiontophoresis entails movement of ions from a micropipette in response to a difference in electrical potential between the inside and outside of the micropipette. Biologically active molecules are thereby delivered in proportion to electrical current. We illustrate acetylcholine microiontophoresis in conjunction with micromanipulation to study endothelium-dependent vasodilation in the microcirculation. Biology The Mouse Cremaster Muscle Preparation for Intravital Imaging of the Microcirculation Pooneh Bagher1, Steven S. Segal1,2 1Department of Medical Pharmacology and Physiology, University of Missouri, 2Dalton Cardiovascular Research Center, University of Missouri A tissue preparation is described for visualization and experimental manipulation of the living microcirculation. In anesthetized male mice, the thin, highly vascularized cremaster muscle is prepared for intravital microscopy to study microvascular networks including arterioles, capillaries and venules. This preparation is readily adapted for rats and hamsters. Biology An Alternant Method to the Traditional NASA Hindlimb Unloading Model in Mice J. Andries Ferreira1, Jacqueline M. Crissey2, Marybeth Brown1,2 1Physical Therapy Department, University of Missouri, Columbia, 2Biomedical Sciences Department, University of Missouri, Columbia We developed an alternant hindlimb unloading model in mice. The primary advantage of our hindlimb unloading tail-ring method over the conventional Morey-Holton tail-traction technique is a simple straightforward procedure that minimizes stress upon the animal.
