1Department of Biological Engineering, Massachusetts Institute of Technology, 2Environmental Toxicology, Chulabhorn Graduate Institute, 3Department of Biomedical Engineering, University of Minnesota
We describe here a platform that allows comet assay detection of DNA damage with unprecedented throughput. The device patterns mammalian cells into a microarray and enables parallel processing of 96 samples. The approach facilitates analysis of base level DNA damage, exposure-induced DNA damage and DNA repair kinetics.
Published October 18, 2014. Keywords: Bioengineering, comet assay, electrophoresis, microarray, DNA damage, DNA repair
JoVE Clinical and Translational Medicine
1Division of Gastroenterology, BC Children's Hospital
DNBS/TNBS induced colitis offers an alternative and inexpensive method to study the pathobiology of inflammatory bowel disease. The protocol discussed in this paper outlines the successful application of DNBS to induce colitis in mice and rats and allows one to thoroughly study host-mediated intestinal responses.
Published February 27, 2014. Keywords: Medicine, Chemical colitis, Inflammatory Bowel Disease, intra rectal administration, intestinal inflammation, transmural inflammation, myeloperoxidase activity
1Department of Entomology, Rutgers University
A push-pull method for collecting plant volatiles is described. The method allows for a comparison of volatiles induced by herbivore feeding, exogenous methyl jasmonate, and mechanical damage. This technique is also used to investigate the volatile response of undamaged branches to exposure to volatiles from herbivore-damaged branches within blueberry plants.
Published December 18, 2011. Keywords: Plant Biology, herbivore-induced plant volatiles, HIPV, eavesdropping, plant defense, priming
1Department of Anatomy and Cell Biology, Wayne State University School of Medicine, 2Department of Opthalmology, Wayne State University School of Medicine
Multiple light damage protocols have been described to damage photoreceptors and consequently induce a retinal regeneration response in adult zebrafish. This protocol describes an improved method that can be used in pigmented animals and that damages the vast majority of rod and cone photoreceptors across the entire retina.
Published October 24, 2013. Keywords: Neuroscience, Zebrafish, Retinal Degeneration, Retina, Photoreceptor, Müller glia, Light damage
1Heart Research Center Goettingen, 2Clinic of Cardiology & Pulmonology, University Medical Center Goettingen, 3German Center for Cardiovascular Research (DZHK) partner site Goettingen, 4BioMET, Center for Biomedical Engineering & Technology, University of Maryland School of Medicine
In cardiac myocytes, tubular membrane structures form intracellular networks. We describe optimized protocols for i) isolation of myocytes from mouse heart including quality control, ii) live cell staining for state-of-the-art fluorescence microscopy, and iii) direct image analysis to quantify the component complexity and the plasticity of intracellular membrane networks.
Published October 15, 2014. Keywords: Bioengineering, cardiac myocyte, atria, ventricle, heart, primary cell isolation, fluorescence microscopy, membrane tubule, transverse-axial tubule system, image analysis, image processing, T-tubule, collagenase
1Department of Biology, University of North Carolina at Charlotte
Xenopus egg extract is a useful model system to investigate the DNA damage checkpoint. This protocol is for the preparation of Xenopus egg extracts and DNA damage checkpoint inducing reagents. These techniques are adaptable to a variety of DNA damaging approaches in the study of the DNA damage checkpoint signaling.
Published November 5, 2012. Keywords: Genetics, Molecular Biology, Cellular Biology, Developmental Biology, DNA damage checkpoint, Xenopus egg extracts, Xenopus laevis, Chk1 phosphorylation, ATR, AT70, MMS, UV, immunoblotting
1Biotechnology and Cellular Signalling, UMR 7242 CNRS, Université de Strasbourg
We describe a protocol to examine the development of opioid-induced hyperalgesia and tolerance in mice. Based on the measurement of thermal and mechanical nociceptive responses of naïve and morphine-treated animals, it allows to quantify the increase in pain sensitivity (hyperalgesia) and decrease in analgesia (tolerance) associated with chronic opiate administration.
Published July 29, 2014. Keywords: Neuroscience, mice, nociception, tail immersion test, tail pressure test, morphine, analgesia, opioid-induced hyperalgesia, tolerance
1Department of Molecular Biotechnology and Health Science, University of Torino, 2Laboratory of Endothelial Molecular Biology, Vesalius Research Center, VIB
Here we report a protocol to measure oxidative stress in living zebrafish embryos. This procedure allows reactive oxygen species (ROS) detection in both whole embryo tissues and single-cell populations. This protocol will accomplish both qualitative and quantitative analyses.
Published July 7, 2014. Keywords: Developmental Biology, Danio rerio, zebrafish embryos, endothelial cells, redox state analysis, oxidative stress detection, in vivo ROS measurements, FACS (fluorescence activated cell sorter), molecular probes
1Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 2Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, 3Division of Neuropathology, Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 4Curriculum in Genetics and Molecular Biology, University of North Carolina School of Medicine, 5Biological and Biomedical Sciences Program, University of North Carolina School of Medicine, 6Department of Radiation Oncology, Emory University School of Medicine, 7Department of Neurology, Neurosciences Center, University of North Carolina School of Medicine
Phenotypically wild-type astrocytes and neural stem cells harvested from mice engineered with floxed, conditional oncogenic alleles and transformed via viral Cre-mediated recombination can be used to model astrocytoma pathogenesis in vitro and in vivo by orthotopic injection of transformed cells into brains of syngeneic, immune-competent littermates.
Published August 12, 2014. Keywords: Neuroscience, astrocytoma, cortical astrocytes, genetically engineered mice, glioblastoma, neural stem cells, orthotopic allograft
JoVE Clinical and Translational Medicine
1Department of Emergency Medicine, Wayne State University School of Medicine, 2Cardiovascular Research Institute, Wayne State University School of Medicine, 3Department of Physiology, Wayne State University School of Medicine
Bilateral carotid occlusion coupled with systemic hypotension produces global brain ischemia in the rat, resulting in damage to the hippocampus with reproducible severity. Animal subjects are impaired with predictable patterns of brain damage, they recover expediently, and mortality rates are comparatively low.
Published June 22, 2013. Keywords: Medicine, Biomedical Engineering, Neurobiology, Neuroscience, Immunology, Anatomy, Physiology, Cardiology, Brain Ischemia, ischemia, reperfusion, cardiac arrest, resuscitation, 2VOH, brain injury model, CA1 hippocampal neurons, brain, neuron, blood vessel, occlusion, hypotension, animal model