University of Chicago View Institution's Website 47 articles published in JoVE Bioengineering Using Vertically Aligned Carbon Nanofiber Arrays on Rigid or Flexible Substrates for Delivery of Biomolecules and Dyes to Plants Jessica M. Morgan1, Joanna Jelenska2, Dale K. Hensley3, Pengju Li4, Bernadeta R. Srijanto3, Scott T. Retterer3,5, Robert F. Standaert6, Jennifer L. Morrell-Falvey5, Jean T. Greenberg2 1Biophysical Sciences, The University of Chicago, 2Molecular Genetics and Cell Biology, The University of Chicago, 3Center for Nanophase Materials Sciences, Oak Ridge National Laboratory, 4Pritzker School of Molecular Engineering, The University of Chicago, 5Biosciences Division, Oak Ridge National Laboratory, 6Department of Chemistry, East Tennessee State University Here we describe methods for microfabricating vertically aligned carbon nanofibers (VACNFs), transferring VACNFs to flexible substrates, and applying VACNFs on both rigid and flexible substrates to plants for biomolecule and dye delivery. Biology Characterizing Epithelial Wound Healing In Vivo Using the Cnidarian Model Organism Clytia hemisphaerica Elizabeth E. L. Lee1, Emily Watto1, Jocelyn Malamy1 1Department of Molecular Genetics and Cell Biology, The University of Chicago This paper describes a method to create wounds in the epithelium of a live Clytia hemisphaerica medusa and image wound healing at a high resolution in vivo. Additionally, a technique to introduce dyes and drugs to perturb signaling processes in the epithelial cells and extracellular matrix during wound healing is presented. Cancer Research Far-Red Fluorescent Senescence-Associated β-Galactosidase Probe for Identification and Enrichment of Senescent Tumor Cells by Flow Cytometry Amy Flor1, Joanna Pagacz1, DeShawn Thompson1, Stephen Kron1 1Department of Molecular Genetics and Cell Biology, University of Chicago A protocol for fluorescent, flow cytometric quantification of senescent cancer cells induced by chemotherapy drugs in cell culture or in murine tumor models is presented. Optional procedures include co-immunostaining, sample fixation to facilitate large batch or time point analysis, and the enrichment of viable senescent cells by flow cytometric sorting. Developmental Biology Eye Removal in Living Zebrafish Larvae to Examine Innervation-dependent Growth and Development of the Visual System Olivia L. Hagen1, Yehyun Kim1, Elaine Kushkowski1,2, Hannah Rouse3, Kara L. Cerveny1 1Department of Biology, Reed College, 2Committee on Development, Regeneration, and Stem Cell Biology, University of Chicago, 3Department of Cell and Developmental Biology, University College London The article explains how to surgically remove eyes from living zebrafish larvae as the first step toward investigating how retinal input influences optic tectum growth and development. In addition, the article provides information about larval anesthetization, fixation, and brain dissection, followed by immunohistochemistry and confocal imaging. Bioengineering An In vitro System to Gauge the Thrombolytic Efficacy of Histotripsy and a Lytic Drug Aarushi Bhargava1, Samuel A. Hendley2, Kenneth B. Bader1,2 1Department of Radiology, University of Chicago, 2Graduate Program in Medical Physics, University of Chicago Histotripsy-aided lytic delivery or lysotripsy is under development for the treatment of deep vein thrombosis. An in vitro procedure is presented here to assess the efficacy of this combination therapy. Key protocols for the clot model, image guidance, and assessment of treatment efficacy are discussed. Engineering Building an Enhanced Flight Mill for the Study of Tethered Insect Flight Anastasia Bernat1 1Department of Ecology and Evolution, University of Chicago This protocol uses three-dimensional (3D) printers and laser cutters found in makerspaces in order to create a more flexible flight mill design. By using this technology, researchers can reduce costs, enhance design flexibility, and generate reproducible work when constructing their flight mills for tethered insect flight studies. Bioengineering Silicon Nanowires and Optical Stimulation for Investigations of Intra- and Intercellular Electrical Coupling Menahem Y. Rotenberg*1, Erik N. Schaumann*2, Aleksander Prominski1,2, Bozhi Tian1,2,3 1The James Franck Institute, The University of Chicago, 2Department of Chemistry, The University of Chicago, 3The Institute for Biophysical Dynamics, The University of Chicago This protocol describes the use of silicon nanowires for intracellular optical bio-modulation of cell in a simple and easy to perform method. The technique is highly adaptable to diverse cell types and can be used for in vitro as well as in vivo applications. Chemistry An Externally-Heated Diamond Anvil Cell for Synthesis and Single-Crystal Elasticity Determination of Ice-VII at High Pressure-Temperature Conditions Xiaojing Lai1,2, Feng Zhu2, Jin S. Zhang3, Dongzhou Zhang2,4, Sergey Tkachev4, Vitali B. Prakapenka4, Bin Chen2 1Gemmological Institute, China University of Geosciences (Wuhan), 2Hawai'i Institute of Geophysics and Planetology, University of Hawai'i at Mānoa, 3Department of Earth and Planetary Sciences, University of New Mexico, 4Center for Advanced Radiation Sources, University of Chicago This work focuses on the standard protocol for preparing the externally-heated diamond anvil cell (EHDAC) for generating high-pressure and high-temperature (HPHT) conditions. The EHDAC is employed to investigate materials in Earth and planetary interiors under extreme conditions, which can be also used in solid state physics and chemistry studies. Chemistry Assembly and Characterization of Polyelectrolyte Complex Micelles Alexander E. Marras1, Jeffrey R. Vieregg1, Matthew V. Tirrell1 1Pritzker School of Molecular Engineering, The University of Chicago We provide protocols and representative data for designing, assembling, and characterizing polyelectrolyte complex micelles, core-shell nanoparticles formed by polyelectrolytes and hydrophilic charged-uncharged block copolymers. Biology Quantification of Proliferative and Dead Cells in Enteroids Hua-Shan Li*1, Shao-Fang Xu*1, Jian-Ying Sheng*1, Zhi-Hui Jiang1, Jing Wang1, Ning Ding1, Tao Wang1, Matthew A. Odenwald2, Jerrold R. Turner2,3, Wei-Qi He1, Hong Xu1, Juan-Min Zha1 1Jiangsu Key Laboratory of Neuropsychiatric Diseases and Cambridge-Suda (CAM-SU) Genomic Resource Center, Medical College of Soochow University, Department of Oncology, The First Affiliated Hospital of Soochow University, 2Department of Pathology, University of Chicago, 3 The presented protocol uses flow cytometry to quantify the number of proliferating and dead cells in cultured mouse enteroids. This method is helpful to evaluate the effects of drug treatment on organoid proliferation and survival. Biochemistry Visualizing Surface T-Cell Receptor Dynamics Four-Dimensionally Using Lattice Light-Sheet Microscopy Jillian Rosenberg1, Jun Huang1,2 1Committee on Cancer Biology, The University of Chicago, 2Pritzker School of Molecular Engineering, The University of Chicago The goal of this protocol is to show how to use Lattice Light-Sheet Microscopy to four-dimensionally visualize surface receptor dynamics in live cells. Here T cell receptors on CD4+ primary T cells are shown. Engineering Procedure for the Transfer of Polymer Films Onto Porous Substrates with Minimized Defects Lorenzo Guio*1, Claire Liu*1, Dean Boures*1, Patrick T. Getty*1, Ruben Waldman1,2, Xiaoying Liu1, Seth B. Darling1,2,3,4 1Institute for Molecular Engineering, University of Chicago, 2Chemical Sciences and Engineering Division, Argonne National Laboratory, 3Institute for Molecular Engineering, Argonne National Laboratory, 4Advanced Materials for Energy-Water Systems (AMEWS) Energy Frontier Research Center, Argonne National Laboratory We present a procedure for highly controlled and wrinkle-free transfer of block copolymer thin films onto porous support substrates using a 3D-printed drain chamber. The drain chamber design is of general relevance to all procedures involving transfer of macromolecular films onto porous substrates, which is normally done by hand in an irreproducible fashion. Biology 4D Microscopy of Yeast Natalie Johnson1, Benjamin S. Glick1 1Department of Molecular Genetics and Cell Biology, University of Chicago This protocol describes the analysis of fluorescently labeled intracellular compartments in budding yeast using multi-color 4D (time-lapse 3D) confocal microscopy. The imaging parameters are chosen to capture adequate signals while limiting photodamage. Custom ImageJ plugins allow labeled structures to be tracked and quantitatively analyzed. Neuroscience Real-time Video Projection in an MRI for Characterization of Neural Correlates Associated with Mirror Therapy for Phantom Limb Pain Faddi G. Saleh Velez1,2, Camila B. Pinto1,3, Emma S. Bailin4, Marionna Münger1, Andrew Ellison5, Beatriz T. Costa1, David Crandell6, Nadia Bolognini7,8, Lotfi B. Merabet4, Felipe Fregni1 1Laboratory of Neuromodulation & Center for Clinical Research Learning, Department of Physical Medicine and Rehabilitation, Harvard Medical School, Spaulding Rehabilitation Hospital, 2University of Chicago Medical Center, Department of Neurology, University of Chicago, 3Department of Neuroscience and Behavior, Psychology Institute, University of Sao Paulo, 4The Laboratory for Visual Neuroplasticity, Department of Ophthalmology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, 5Center for Biomedical Imaging, Department of Anatomy and Neurobiology, Boston University School of Medicine, 6Spaulding Rehabilitation Hospital, Harvard Medical School, 7Department of Psychology & Milan Center for Neuroscience, University of Milano-Bicocca, 8Neuropsychological Laboratory, Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Istituto Auxologico Italiano We present a novel combined behavioral and neuroimaging protocol employing real-time video projection for the purpose of characterizing the neural correlates associated with mirror therapy within the magnetic resonance imaging scanner environment in leg amputee subjects with phantom limb pain. Bioengineering Conducting Multiple Imaging Modes with One Fluorescence Microscope Seongjin Park1, Jiacheng Zhang2, Matthew A. Reyer2, Joanna Zareba1,3, Andrew A. Troy4, Jingyi Fei1,2 1Department of Biochemistry and Molecular Biology, University of Chicago, 2The Institute for Biophysical Dynamics, University of Chicago, 3Faculty of Chemistry, Wrocław University of Science and Technology, 4Nikon Instruments Inc. Here we present a practical guide of building an integrated microscopy system, which merges conventional epi-fluorescent imaging, single-molecule detection-based super-resolution imaging, and multi-color single-molecule detection, including single-molecule fluorescence resonance energy transfer imaging, into one set-up in a cost-efficient way. Immunology and Infection Generation of Escape Variants of Neutralizing Influenza Virus Monoclonal Antibodies Paul E. Leon1,2, Teddy John Wohlbold1,2, Wenqian He1,2, Mark J. Bailey1,2, Carole J. Henry3, Patrick C. Wilson3, Florian Krammer1, Gene S. Tan1 1Department of Microbiology, Icahn School of Medicine at Mount Sinai, 2Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, 3The Department of Medicine, Section of Rheumatology, The Knapp Center for Lupus and Immunology Research, The University of Chicago We describe a method by which we identify critical residues required for the binding of human or murine monoclonal antibodies that target the viral hemagglutinin of influenza A viruses. The protocol can be adapted to other virus surface glycoproteins and their corresponding neutralizing antibodies. Medicine Automated Radiochemical Synthesis of [18F]3F4AP: A Novel PET Tracer for Imaging Demyelinating Diseases Pedro Brugarolas1, Mohammed Bhuiyan2, Anna Kucharski2, Richard Freifelder2 1Department of Neurology, The University of Chicago, 2Department of Radiology, The University of Chicago We demonstrate the semi-automated radiochemical synthesis of [18F]3F4AP and quality control procedures. Immunology and Infection Isolation of the Side Population in Myc-induced T-cell Acute Lymphoblastic Leukemia in Zebrafish Margaret M. Pruitt1,2, Wilfredo Marin1, Michael R. Waarts1, Jill L. O. de Jong1 1Department of Pediatrics, Section of Hematology-Oncology, The University of Chicago, 2Department of Biology and Biotechnology, Worcester Polytechnic Institute Here, we describe a technique to isolate the side population cells from a zebrafish model of myc-induced T-cell acute lymphoblastic leukemia (T-ALL). This side population assay is highly sensitive and is described for zebrafish T-ALL, but it may be applicable to other malignant and non-malignant zebrafish cell types. Chemistry High Pressure Single Crystal Diffraction at PX^2 Dongzhou Zhang1, Przemyslaw K. Dera1, Peter J. Eng2, Joanne E. Stubbs2, Jin S. Zhang1, Vitali B. Prakapenka2, Mark L. Rivers2 1Hawai’i Institute of Geophysics and Planetology, University of Hawai’i at Manoa, 2Center for Advanced Radiation Sources, University of Chicago In this report, we describe detailed procedures for carrying out single crystal X-ray diffraction experiments with a diamond anvil cell at the GSECARS 13-BM-C beamline at the Advanced Photon Source. ATREX and RSV programs are used to analyze the data. Cancer Research Advanced Animal Model of Colorectal Metastasis in Liver: Imaging Techniques and Properties of Metastatic Clones Go Oshima*1,2, Melinda E. Stack*1, Sean C. Wightman1, Darren Bryan1, Elizabeth Poli1, Lai Xue1, Kinga B. Skowron1, Abhineet Uppal1, Sean P. Pitroda2, Xiaona Huang2, Mitchell C. Posner1, Samuel Hellman2, Ralph R. Weichselbaum2, Nikolai N. Khodarev2 1Department of Surgery, The University of Chicago, 2Department of Radiation and Cellular Oncology and Ludwig Center for Metastasis Research, The University of Chicago The ability of metastatic clones to colonize distant sites depends on their proliferation capacity and/or their ability to survive in the host microenvironment without significant proliferation. Here, we present an animal model that allows quantitative visualization of both types of liver colonization by metastatic clones. Neuroscience Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior Xiao Sun1, Ellie S. Heckscher1,2 1Committee on Development, Regeneration, and Stem Cell Biology, University of Chicago, 2Department of Molecular Genetics and Cell Biology, University of Chicago The Drosophila larva is a powerful model system to study neural control of behavior. This publication describes the use of linear agarose channels to elicit sustained bouts of linear crawling and methods to quantify the dynamics of larval structures during repetitive crawling behavior. Biology Filtration Isolation of Nucleic Acids: A Simple and Rapid DNA Extraction Method Sally M. McFall1, Mário F. Neto1, Jennifer L. Reed1, Robin L. Wagner2 1Center for Innovation in Global Health Technologies (CIGHT), Department of Biomedical Engineering, Northwestern University, 2Pritzker School of Medicine, The University of Chicago We describe here a simple and rapid paper-based DNA extraction method of HIV proviral DNA from whole blood detected by quantitative PCR. This protocol can be extended for use in detecting other genetic markers or using alternative amplification methods. Neuroscience Quantification of Cerebral Vascular Architecture using Two-photon Microscopy in a Mouse Model of HIV-induced Neuroinflammation Christopher Nishimura1, Oksana Polesskaya2, Stephen Dewhurst1, Jharon N. Silva1 1Department of Microbiology and Immunology, University of Rochester Medical Center, 2Department of Human Genetics, University of Chicago This paper describes a method by which the vascular architecture in the brain can be quantified using in vivo and ex vivo two-photon microscopy. Medicine Modeling the Early Steps of Ovarian Cancer Dissemination in an Organotypic Culture of the Human Peritoneal Cavity Pamela N. Peters1, Elizabeth M. Schryver1, Ernst Lengyel1, Hilary Kenny1 1Department of Obstetrics and Gynecology, Section of Gynecologic Oncology, University of Chicago Here, we present a protocol to construct a three-dimensional in vitro model of the lining of the peritoneal cavity, composed of primary human mesothelial cells and fibroblasts layered with extracellular matrix, as a tool to investigate ovarian cancer cell adhesion, invasion, and proliferation. Medicine In Vivo and Ex Vivo Approaches to Study Ovarian Cancer Metastatic Colonization of Milky Spot Structures in Peritoneal Adipose Venkatesh Krishnan1, Robert Clark1, Marina Chekmareva2, Amy Johnson1, Sophia George3, Patricia Shaw4, Victoria Seewaldt4,5, Carrie Rinker-Schaeffer1 1Section of Urology, Department of Surgery, The University of Chicago, 2Department of Pathology, Robert Wood Johnson Medical School, 3Campbell Family Institute for Breast Cancer Research, Princess Margaret Cancer Centre, University Health Network, 4Department of Laboratory Medicine and Pathobiology, Campbell Family Institute for Breast Cancer Research, Princess Margaret Cancer Centre, University of Toronto, University Health Network, 5Departments of Medicine, Pharmacology, and Cancer Biology, Duke University Medical Center We outline a protocol that implements both in vivo and ex vivo approaches to study ovarian cancer colonization of peritoneal adipose tissues, particularly the omentum. Furthermore, we present a protocol to quantitate and analyze immune cell-structures in the omentum known as milky spots, which promote metastases of peritoneal adipose. Biology Surface Spreading and Immunostaining of Yeast Chromosomes Jennifer Grubb1, M. Scott Brown1,2, Douglas K. Bishop1,2 1Department of Radiation and Cellular Oncology, University of Chicago, 2Department of Molecular Genetics and Cell Biology, University of Chicago A method for surface-spreading chromosomes from budding yeast is presented. This method is derived from a method previously described by Loidl and Klein. In addition, we demonstrate a procedure for immunostaining of spread chromosomes. Immunology and Infection Scalable High Throughput Selection From Phage-displayed Synthetic Antibody Libraries Shane Miersch1,2, Zhijian Li1,2, Rachel Hanna1,2, Megan E. McLaughlin1,2, Michael Hornsby1,3, Tet Matsuguchi1,3, Marcin Paduch1,4, Annika Sääf1,4, Jim Wells1,3, Shohei Koide1,4, Anthony Kossiakoff1,4, Sachdev S. Sidhu1,2 1The Recombinant Antibody Network, 2The Banting and Best Department of Medical Research, University of Toronto, 3Antibiome Center, University of California, San Francisco at Mission Bay, 4Department of Biochemistry and Molecular Biology, The University of Chicago A method is described with visual accompaniment for conducting scalable, high throughput selections from phage-displayed combinatorial synthetic antibody libraries against hundreds of antigens simultaneously. Using this parallel approach, we have isolated antibody fragments that exhibit high affinity and specificity for diverse antigens that are functional in standard immunoassays. Engineering Fast Imaging Technique to Study Drop Impact Dynamics of Non-Newtonian Fluids Qin Xu1,2, Ivo Peters2, Sam Wilken1,2, Eric Brown3, Heinrich Jaeger1,2 1Department of Physics, The University of Chicago, 2James Franck Institute, The University of Chicago, 3Department of Mechanical Engineering and Materials Science, Yale University Drop impact of non-Newtonian fluids is a complex process since different physical parameters influence the dynamics over a very short time (less than one tenth of a millisecond). A fast imaging technique is introduced in order to characterize the impact behaviors of different non-Newtonian fluids. Neuroscience Laser-scanning Photostimulation of Optogenetically Targeted Forebrain Circuits Charles C. Lee1, Ying-Wan Lam2, Kazuo Imaizumi1, S. Murray Sherman2 1Department of Comparative Biomedical Sciences, School of Veterinary Medicine, Louisiana State University, 2Department of Neurobiology, University of Chicago We describe a method for characterizing the functional topography and synaptic properties of forebrain circuits using an optogenetic approach to photostimulate neuronal populations in vitro. Chemistry Seeded Synthesis of CdSe/CdS Rod and Tetrapod Nanocrystals Karthish Manthiram*1,4, Brandon J. Beberwyck*2,4, Dmitri V. Talapin5,6, A. Paul Alivisatos2,3,4 1Department of Chemical Engineering, UC Berkeley, 2Department of Materials Science and Engineering, UC Berkeley, 3Department of Chemistry, UC Berkeley, 4Materials Sciences Division, Lawrence Berkeley National Laboratory, 5Department of Chemistry, University of Chicago, 6Center for Nanoscale Materials, Argonne National Laboratory A protocol for the seeded synthesis of rod-shaped and tetrapod-shaped multicomponent nanostructures consisting of CdS and CdSe is presented. Engineering Synthesis and Microdiffraction at Extreme Pressures and Temperatures Barbara Lavina1, Przemyslaw Dera2, Yue Meng3 1High Pressure Science and Engineering Center, Department of Physics and Astronomy, University of Nevada, Las Vegas, 2GeoSoilEnviroCARS, University of Chicago, 3High Pressure Collaborative Access Team, Carnegie Institution of Washington The laser heated diamond anvil cell combined with synchrotron micro-diffraction techniques allows researchers to explore the nature and properties of new phases of matter at extreme pressure and temperature (PT) conditions. Heterogeneous samples can be characterized in situ under high pressure by 2D mapping and combined powder, single-crystal and multigrain diffraction approaches. Medicine Formation of Human Prostate Epithelium Using Tissue Recombination of Rodent Urogenital Sinus Mesenchyme and Human Stem Cells Yi Cai1, Steven Kregel2, Donald J. Vander Griend1,2 1Department of Surgery, Section of Urology, University of Chicago, 2Committee on Cancer Biology, University of Chicago To unravel the earliest molecular mechanisms underlying prostate cancer initiation, novel and innovative human model systems and approaches are desperately needed. The potential of pre-prostatic urogenital sinus mesenchyme (UGSM) to induce pluripotent stem cell populations to form human prostate epithelium is a powerful experimental tool in prostate research. Medicine Murine Fetal Echocardiography Gene H. Kim1 1Department of Medicine, Section of Cardiology, Institute for Cardiovascular Research, University of Chicago Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development. High-frequency ultrasound imaging has improved 2-D resolution and can provide excellent information on early cardiac development and is an ideal method to detect the impact on cardiac structure and function prior to death. Engineering Concurrent Quantitative Conductivity and Mechanical Properties Measurements of Organic Photovoltaic Materials using AFM Maxim P. Nikiforov1, Seth B. Darling1,2 1Center for Nanoscale Materials, Argonne National Laboratory, 2Institute for Molecular Engineering, University of Chicago Organic photovoltaic (OPV) materials are inherently inhomogeneous at the nanometer scale. Nanoscale inhomogeneity of OPV materials affects performance of photovoltaic devices. In this paper, we describe a protocol for quantitative measurements of electrical and mechanical properties of OPV materials with sub-100 nm resolution. Biology Selective Capture of 5-hydroxymethylcytosine from Genomic DNA Yujing Li1, Chun-Xiao Song2, Chuan He2, Peng Jin1 1Department of Human Genetics, Emory University School of Medicine, 2Department of Chemistry and Institute for Biophysical Dynamics, The University of Chicago Described is a two-step labeling process using β-glucosyltransferase (β-GT) to transfer an azide-glucose to 5-hmC, followed by click chemistry to transfer a biotin linker for easy and density-independent enrichment. This efficient and specific labeling method enables enrichment of 5-hmC with extremely low background and high-throughput epigenomic mapping via next-generation sequencing. Neuroscience Modeling Neural Immune Signaling of Episodic and Chronic Migraine Using Spreading Depression In Vitro Aya D. Pusic*1, Yelena Y. Grinberg*1, Heidi M. Mitchell2, Richard P. Kraig1 1Department of Neurology and Committee on Neurobiology, The University of Chicago Medical Center, 2Department of Neurology, The University of Chicago Medical Center Migraine and its transformation to chronic migraine are immense healthcare burdens in need of improved treatment options. We seek to define how neural immune signaling modulates the susceptibility to migraine, modeled in vitro using spreading depression in hippocampal slice cultures, as a means to develop novel therapeutic targets. Biology Computer-assisted Large-scale Visualization and Quantification of Pancreatic Islet Mass, Size Distribution and Architecture Abraham Kim1, German Kilimnik1, Charles Guo1, Joshua Sung1, Junghyo Jo2, Vipul Periwal2, Piotr Witkowski3, Philip Dilorio4, Manami Hara1 1Department of Medicine, University of Chicago, 2Laboratory of Biological Modeling, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 3Department of Surgery, University of Chicago, 4Diabetes Division, University of Massachusetts Novel computer-assisted methods of large-scale procurement and analysis of immunohistochemically stained pancreatic specimens are described: (1) Virtual Slice capture of the entire section; (2) Mass analysis of large-scale data; (3) Reconstruction of 2D Virtual Slices; (4) 3D islet mapping; and (5) Mathematical analysis. Medicine Generation of Comprehensive Thoracic Oncology Database - Tool for Translational Research Mosmi Surati1, Matthew Robinson2, Suvobroto Nandi2, Leonardo Faoro2, Carley Demchuk2, Rajani Kanteti2, Benjamin Ferguson1, Tara Gangadhar2, Thomas Hensing3, Rifat Hasina2, Aliya Husain4, Mark Ferguson5, Theodore Karrison6, Ravi Salgia2 1Pritzker School of Medicine, University of Chicago, 2Department of Medicine, University of Chicago, 3Department of Medicine, Northshore University Health Systems, 4Department of Pathology, University of Chicago, 5Department of Surgery, University of Chicago, 6Department of Biostatistics, University of Chicago A thoracic oncology database was developed to serve as a comprehensive repository for clinical and laboratory data for the purposes of translational research. The database will serve translational cancer researchers within the Thoracic Oncology Research Program. This database is adaptable to other cancer models, as well as other human diseases. Neuroscience Combining Computer Game-Based Behavioural Experiments With High-Density EEG and Infrared Gaze Tracking Keith J. Yoder1,2, Matthew K. Belmonte1,3 1Department of Human Development, Cornell University, 2Social Sciences Division, University of Chicago, 3National Brain Research Centre, Manesar, India Procedures for recording high-density EEG and gaze data during computer game-based cognitive tasks are described. Using a video game to present cognitive tasks enhances ecological validity without sacrificing experimental control. Bioengineering Preparation of Complaint Matrices for Quantifying Cellular Contraction Yvonne Aratyn-Schaus1, Patrick W. Oakes1, Jonathan Stricker2, Stephen P. Winter3, Margaret L. Gardel1,2 1Institute for Biophysical Dynamics, University of Chicago, 2Physics Department - James Franck Institute, University of Chicago, 3Interdisciplinary Scientist Training Program, University of Chicago In this video, we demonstrate the experimental techniques used to fabricate compliant, extracellular matrix (ECM) coated substrates suitable for cell culture, and which are amenable to traction force microscopy and observing effects of ECM stiffness on cell behavior. Neuroscience Strategies for Study of Neuroprotection from Cold-preconditioning Heidi M. Mitchell1, David M. White1, Richard P. Kraig1 1Department of Neurology, The University of Chicago Medical Center We seek to define the neural immune signaling responsible for cold-preconditioning as means to identify novel targets for therapeutics development to protect brain before injury onset. We present strategies for such work that require biological systems, experimental manipulations plus technical capacities that are highly reproducible and sensitive. Biology Intraperitoneal Injection into Adult Zebrafish Mary D. Kinkel1, Stefani C. Eames2, Louis H. Philipson2,3, Victoria E. Prince1 1Department of Organismal Biology and Anatomy, The University of Chicago, 2Committee on Molecular Metabolism and Nutrition, The University of Chicago, 3Department of Medicine, The University of Chicago We demonstrate intraperitoneal injection into adult zebrafish. We use a 10 μl NanoFil microsyringe controlled by a Micro4 controller and UltraMicroPump III. This demonstration includes the use of cold water as an anesthetic. Biology Genome-wide Analysis of Aminoacylation (Charging) Levels of tRNA Using Microarrays John Zaborske1, Tao Pan1 1Department of Biochemistry and Molecular Biology, University of Chicago We describe a method for microarray analysis to determine relative aminoacylation levels of all tRNAs from S. cerivisiae. Biology In situ Quantification of Pancreatic Beta-cell Mass in Mice Abraham Kim1, German Kilimnik1, Manami Hara1 1Department of Medicine, University of Chicago The following protocol outlines the process of pancreatic dissection for virtual slice imaging, and the subsequent quantification of all GFP-tagged beta-cells in the entire pancreas. Application Notes The NanoDrop 2000c Spectrophotometer - Microvolume Protein Concentration Determination (Spanish) - ADVERTISEMENT Theodore Karrison1 1Department of Health Studies, University of Chicago Biology In Ovo Electroporations of HH Stage 10 Chicken Embryos Marissa C. Blank1, Victor Chizhikov2, Kathleen J. Millen2 1Department of Molecular Genetics and Cell Biology, University of Chicago, 2Department of Human Genetics, University of Chicago Chick in ovo electroporation is a technique which allows genetic manipulation of the avian embryo. Common applications of this technique include functional analysis of genes and putative enhancer elements. This video demonstrates neural tube electroporation in HH 10 chick embryos. Injection technique and proper egg handling are discussed. Biology Understanding Cerebellar Pattern Formation Kathleen J. Millen1 1Department of Neurology, University of Chicago