Universidad Complutense de Madrid 11 articles published in JoVE Neuroscience Cell-Type Specific Protein Purification and Identification from Complex Tissues Using a Mutant Methionine tRNA Synthetase Mouse Line Belquis Nassim-Assir1, Daniel Ouro Corredera2, Rodrigo Alvarez-Pardo2, Susanne tom Dieck1, Elena Ciirdaeva1, Erin M. Schuman1, Beatriz Alvarez-Castelao2 1Max-Planck-Institute for Brain Research, Synaptic Plasticity Department, 2Department of Biochemistry and Molecular Biology, Veterinary School, Complutense University of Madrid This protocol describes how to perform cell-type-specific protein labeling with azidonorleucine (ANL) using a mouse line expressing a mutant L274G-Methionine tRNA synthetase (MetRS*) and the necessary steps for labeled cell-type-specific proteins isolation. We outline two possible ANL administration routes in live mice by (1) drinking water and (2) intraperitoneal injections. Environment Laboratory and Field Protocol for Estimating Sheet Erosion Rates from Dendrogeomorphology Jose Maria Bodoque1, Juan Antonio Ballesteros-Cánovas2,3, Juan Manuel Rubiales4,5, Markus Stoffel2,3 1University of Castilla-La Mancha (UCLM), 2Department of Earth Sciences, University of Geneva, 3Institute for Environmental Sciences, University of Geneva, 4Departamento de Sistemas y Recursos Naturales, Universidad Politécnica de Madrid, 5Departamento de Biodiversidad, Ecología y Evolución, Universidad Complutense de Madrid Characterizing erosion from dendrogeomorphology has usually focused on accurately finding the starting time of root exposure, by examining macroscopic or cell level changes caused by exposure. Here, we offer a detailed description of different novel techniques to obtain more precise erosion rates from highly accurate microtopographic data. Biology Synthesis of 68Ga Core-doped Iron Oxide Nanoparticles for Dual Positron Emission Tomography /(T1)Magnetic Resonance Imaging Irene Fernández-Barahona1, Jesús Ruiz-Cabello2, Fernando Herranz1, Juan Pellico3 1Nanobiotechnology, Molecular Imaging and Metabolomics Lab, Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), 2CIC biomaGUNE and CIBER de Enfermedades Respiratorias (CIBERES). Ikerbasque, Basque Foundation for Science, Universidad Complutense de Madrid (UCM), 3Advanced Imaging Unit, Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), CIBER de Enfermedades Respiratorias (CIBERES) Here, we present a protocol to obtain 68Ga core-doped iron oxide nanoparticles via fast microwave-driven synthesis. The methodology renders PET/(T1)MRI nanoparticles with radiolabeling efficiencies higher than 90% and radiochemical purity of 99% in a 20-min synthesis. Genetics Testing the Role of Multicopy Plasmids in the Evolution of Antibiotic Resistance Jose Antonio Escudero1,2, R Craig MacLean3, Alvaro San Millan4 1Department of Animal Health, Universidad Complutense de Madrid, 2Visavet Health Surveillance Centre, Universidad Complutense de Madrid, 3Department of Zoology, University of Oxford, 4Department of Microbiology, Hospital Universitario Ramon y Cajal (IRYCIS) and CIBERESP Here we present an experimental method to test the role of multicopy plasmids in the evolution of antibiotic resistance. Engineering Growth and Electrostatic/chemical Properties of Metal/LaAlO3/SrTiO3 Heterostructures Diogo Castro Vaz1, Edouard Lesne1,2, Anke Sander1, Hiroshi Naganuma1,3, Eric Jacquet1, Jacobo Santamaria1,4, Agnès Barthélémy1, Manuel Bibes1 1Unité Mixte de Physique CNRS/Thales, Université Paris-Saclay, 2Max Planck Institut für Mikrostrukturphysik, 3Department of Applied Physics, Tohoku University, 4Instituto de Magnetismo Aplicado, Universidad Complutense de Madrid We fabricate metal/LaAlO3/SrTiO3 heterostructures using a combination of pulsed laser deposition and in situ magnetron sputtering. Through magnetotransport and in situ X-ray photoelectron spectroscopy experiments, we investigate the interplay between electrostatic and chemical phenomena of the quasi two-dimensional electron gas formed in this system. Developmental Biology Bottlenose Dolphin (Tursiops truncatus) Spermatozoa: Collection, Cryopreservation, and Heterologous In Vitro Fertilization María Jesús Sánchez-Calabuig1,2, Francisco Alberto García-Vázquez3, Ricardo Laguna-Barraza1, Carlos Barros-García4, Daniel García-Parraga5, Dimitrios Rizos1, Alfonso Gutiérrez Adan1, José Félix Pérez-Gutíerrez2 1Department of Animal Reproduction, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), 2Department of Animal Medicine and Surgery, School of Veterinary Medicine, Universidad Complutense de Madrid, 3Department of Physiology, Faculty of Veterinary Science, University of Murcia, Campus Mare Nostrum, 4Mundomar, Benidorm, 5 Here, we present protocols that have been successfully used for dolphin spermatozoa collection, cryopreservation, and heterologous IVF performance using bovine oocytes. Immunology and Infection Methodology for the Study of Horizontal Gene Transfer in Staphylococcus aureus Fabio Cafini*1,2, Nguyen Thi Le Thuy*3, Federico Román4, José Prieto5, Sarah Dubrac6,7, Tarek Msadek6,7, Kazuya Morikawa1 1Division of Biomedical Science, Faculty of Medicine, University of Tsukuba, 2Department of Basic Biomedical Science, Universidad Europea de Madrid, 3Human Biology Program, School of Integrative and Global Majors, University of Tsukuba, 4Laboratory of Nosocomial Infections, Department of Bacteriology, Centro Nacional de MicrobiologÍa, Instituto de Salud Carlos III, 5Division of Microbiology, Department of Medicine, School of Medicine, Universidad Complutense, 6Biology of Gram-Positive Pathogens, Department of Microbiology, Institut Pasteur, Paris, France, 7ERL3526, CNRS, Paris, France We describe here three different protocols for the in vitro investigation of conjugation, transduction, and natural transformation in Staphylococcus aureus. Biology A Comparative Study of Drug Delivery Methods Targeted to the Mouse Inner Ear: Bullostomy Versus Transtympanic Injection Silvia Murillo-Cuesta*1,2,3, Néstor Vallecillo*1, Rafael Cediel1,2,4, Adelaida M. Celaya1,2, Luis Lassaletta3,5, Isabel Varela-Nieto1,2,3, Julio Contreras1,2,4 1Instituto de Investigaciones Biomédicas (IIBm) Alberto Sols CSIC-UAM, 2Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Instituto de Salud Carlos III (ISCIII), 3Instituto de Investigación Sanitaria La Paz (IdiPAZ), 4Facultad de Veterinaria, Universidad Complutense de Madrid, 5Departmento de Otorrino laringología, Hospital Universitario La Paz Two microsurgery approaches for local drug delivery to the inner ear are described here and compared in terms of impact on hearing parameters, cochlear cytoarchitecture and expression of inflammatory markers. Immunology and Infection Phenotypic Characterization of Macrophages from Rat Kidney by Flow Cytometry Alfonso Rubio-Navarro1, Melania Guerrero-Hue1, Beatriz Martín-Fernandez2, Isabel Cortegano3, Elena Olivares-Alvaro2, Natalia de las Heras2, Mario Alía3, Belén de Andrés3, María Luisa Gaspar3, Jesús Egido1, Juan Antonio Moreno1 1Renal, Vascular and Diabetes Research Lab, IIS-Fundaciòn Jiménez Dìaz, Autonoma University, 2Department of Physiology, Faculty of Medicine, Complutense University, 3Department of Immunology, Centro Nacional de Microbiologìa, Instituto de Salud Carlos III (ISCIII) This manuscript describes a detailed protocol for phenotypic and quantitative analysis of resident macrophages from rat kidneys by flow cytometry. The resulting stained cells can be also used for other applications, including cell sorting, gene expression analysis or functional studies, thus increasing the information obtained in the experimental model. Bioengineering Microwave-driven Synthesis of Iron Oxide Nanoparticles for Fast Detection of Atherosclerosis Juan Pellico1,2, Jesús Ruiz-Cabello1,2, Fernando Herranz1 1Advanced Imaging Unit, Centro Nacional de Investigaciones Cardiovasculares Carlos III and CIBER de Enfermedades Respiratorias, 2Universidad Complutense de Madrid Microwave technology enables extremely fast synthesis of iron oxide nanoparticles for atherosclerosis plaque characterization. The use of an aminobisphosphonate in the external side of the nanoparticle provides a fast accumulation in the atherosclerotic area. Medicine Stereological and Flow Cytometry Characterization of Leukocyte Subpopulations in Models of Transient or Permanent Cerebral Ischemia Iván Ballesteros*1, María Isabel Cuartero*1, Ana Moraga1, Juan de la Parra1, Ignacio Lizasoain1, María Ángeles Moro1 1Unidad de Investigacón Neurovascular, Departmento de Farmacología, Falcultad de Medicina, Universidad Complutense de Madrid y Instituto de Investigación Hospital 12 de Octubre, Madrid Brain myeloid cells characterization following stroke can be performed by stereology using the optical fractionator method, or by a flow cytometric analysis of brain leukocytes suspensions. Both are useful techniques to perform an accurate phenotypical distinction of the main myeloid cell subsets found in the ischemic brain.