University of California, Davis View Institution's Website 83 articles published in JoVE Genetics High-Efficiency Gene Disruption in Primary Bone Marrow-Derived Macrophages Using Electroporated Cas9-sgRNA Complexes Julia Craft1, Tina Truong1, Bennett H. Penn1,2 1Department of Internal Medicine, University of California, Davis, 2Department of Medical Microbiology and Immunology, University of California, Davis This protocol describes the procedure for genome editing in mouse bone marrow-derived macrophages using Cas9-sgRNA ribonucleoprotein complexes assembled in vitro and delivered by electroporation. Biology Isolation of Small Preantral Follicles from the Bovine Ovary Using a Combination of Fragmentation, Homogenization, and Serial Filtration Stephanie P. McDonnell1, Juliana I. Candelaria1, Amanda J. Morton1, Anna C. Denicol1 1Department of Animal Science, University of California Davis Advancing the study of preantral folliculogenesis requires efficient methods of follicle isolation from single ovaries. Presented here is a streamlined, mechanical protocol for follicle isolation from bovine ovaries using a tissue chopper and homogenizer. This method allows collection of a large number of viable preantral follicles from a single ovary. Genetics Detection of Homologous Recombination Intermediates via Proximity Ligation and Quantitative PCR in Saccharomyces cerevisiae Diedre Reitz1, Jérôme Savocco2, Aurèle Piazza2, Wolf-Dietrich Heyer1,3 1Department of Microbiology & Molecular Genetics, University of California, Davis, 2Laboratory of Biology & Modeling of the Cell, École Normale Supérieure de Lyon, 3Department of Molecular & Cellular Biology, University of California, Davis The D-loop capture (DLC) and D-loop extension (DLE) assays utilize the principle of proximity ligation together with quantitative PCR to quantify D-loop formation, D-loop extension, and product formation at the site of an inducible double-stranded break in Saccharomyces cerevisiae. Biology Systematic Assessment of Mammalian Skull Specimens for Dental and Temporomandibular Joint Pathology Janny Evenhuis1, Boaz Arzi2, Frank J. M. Verstraete2 1Dentistry and Oral Surgery Service, William R. Pritchard Veterinary Medical Teaching Hospital, School of Veterinary Medicine, University of California-Davis, 2Department of Surgical and Radiological Sciences, School of Veterinary Medicine, University of California-Davis The present protocol describes the techniques for the systematic assessment of skull specimens to characterize anatomical and developmental variations and abnormalities of the teeth, periodontal disease, endodontal disease, and temporomandibular joint pathology. Genetics Development of Knock-Out Muscle Cell Lines using Lentivirus-Mediated CRISPR/Cas9 Gene Editing Mathilde Beaufils1, Amandine Tourel1, Anne Petiot1, Nicole B. Halmai2, Dave J. Segal2, John Rendu1, Isabelle Marty1 1University Grenoble Alpes, INSERM, U1216, CHU Grenoble Alpes, Grenoble Institut Neurosciences, 2Genome Center, University of California, Davis The protocol describes how to generate knock-out myoblasts using CRISPR/Cas9, starting from the design of guide-RNAs to the cellular cloning and characterization of the knock-out clones. Neuroscience AAV Deployment of Enhancer-Based Expression Constructs In Vivo in Mouse Brain Tracy L. Warren1,2, Jason T. Lambert1,2, Alex S. Nord1,2 1Department of Psychiatry and Behavioral Sciences, University of California, Davis, 2Department of Neurobiology, Physiology and Behavior, University of California, Davis This protocol describes a novel rAAV-based transient enhancer-reporter assay. This assay can be used to induce enhancer-driven expression in vivo in the mouse brain. Immunology and Infection Cell-Free Scaled Production and Adjuvant Addition to a Recombinant Major Outer Membrane Protein from Chlamydia muridarum for Vaccine Development Sean F. Gilmore*1, Wei He*1, Angela C. Evans1, Delia F. Tifrea2, Sukumar Pal2, Brent Segelke1, Sandra K. G. Peters1, B. Dillon Vannest1, Nicholas O. Fischer1, Amy Rasley1, Luis M. de la Maza2, Matthew A. Coleman1,3 1Physical and Life Sciences Directorate, Lawrence Livermore National Laboratory, 2Department of Pathology and Laboratory Medicine, University of California, 3School of Medicine, Radiation Oncology, University of California Davis This protocol describes using commercial, cell-free protein expression kits to produce membrane proteins supported in nanodisc that can be used as antigens in subunit vaccines. Biology A Magnetic-Bead-Based Mosquito DNA Extraction Protocol for Next-Generation Sequencing Tse-Yu Chen1, Adam E. Vorsino2, Kyle J. Kosinski1, Ana L. Romero-Weaver1, Eva A. Buckner1, Joanna C. Chiu3, Yoosook Lee1 1Florida Medical Entomology Laboratory, Department of Entomology and Nematology, Institute of Food and Agricultural Sciences, University of Florida, 2U. S. Fish and Wildlife Service, Pacific Islands Fish and Wildlife Office, 3Department of Entomology and Nematology, University of California Davis Described here is a DNA extraction protocol using magnetic beads to produce high quality DNA extractions from mosquitoes. These extractions are suitable for a downstream next-generation sequencing approach. Biochemistry Isolation of Histone from Sorghum Leaf Tissue for Top Down Mass Spectrometry Profiling of Potential Epigenetic Markers Mowei Zhou1, Shadan H. Abdali1, David Dilworth2, Lifeng Liu2, Benjamin Cole2, Neha Malhan1, Amir H. Ahkami1, Tanya E. Winkler1, Joy Hollingsworth3, Julie Sievert3, Jeff Dahlberg3, Robert Hutmacher4,5, Mary Madera6, Judith A. Owiti6, Kim K. Hixson1, Peggy G. Lemaux6, Christer Jansson1, Ljiljana Paša-Tolić1 1Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, 2DOE-Joint Genome Institute, Lawrence Berkeley Laboratory, 3Kearney Agricultural Research and Extension Center, University of California Agriculture and Natural Resources, 4West Side Research and Extension Center, University of California, 5Department of Plant Sciences, University of California, Davis, 6Department of Plant and Microbial Biology, University of California, Berkeley The protocol has been developed to effectively extract intact histones from sorghum leaf materials for profiling of histone post-translational modifications that can serve as potential epigenetic markers to aid engineering drought resistant crops. Immunology and Infection TurboID-Based Proximity Labeling for In Planta Identification of Protein-Protein Interaction Networks Yongliang Zhang*1, Yuanyuan Li*2,3, Xinxin Yang1, Zhiyan Wen1, Ugrappa Nagalakshmi2, Savithramma P. Dinesh-Kumar2,3 1State Key Laboratory of Agro-Biotechnology and Ministry of Agriculture Key Laboratory of Soil Microbiology, College of Biological Sciences, China Agricultural University, 2Department of Plant Biology, College of Biological Sciences, University of California, Davis, 3Genome Center, College of Biological Sciences, University of California, Davis Described here is a proximity labeling method for identification of interaction partners of the TIR domain of the NLR immune receptor in Nicotiana benthamiana leaf tissue. Also provided is a detailed protocol for the identification of interactions between other proteins of interest using this technique in Nicotiana and other plant species. Immunology and Infection Identification of Neutrophil Extracellular Traps in Paraffin-Embedded Feline Arterial Thrombi using Immunofluorescence Microscopy Laetitia Duler1, Nghi Nguyen2, Eric Ontiveros3, Ronald H. L. Li2 1Veterinary Medical Teaching Hospital, School of Veterinary Medicine, University of California, Davis, 2Department of Veterinary Surgical and Radiological Sciences, School of Veterinary Medicine, University of California, Davis, 3Department of Veterinary Medicine and Epidemiology, School of Veterinary Medicine, University of California, Davis We describe a method to identify neutrophil extracellular traps (NETs) in formaldehyde-fixed and paraffin-embedded feline cardiogenic arterial thrombi using heat-induced antigen retrieval and a double immunolabeling protocol. Genetics Preparation of Meiotic Chromosome Spreads from Zebrafish Spermatocytes Yana P. Blokhina1,2, Ivan Olaya1,2, Sean M. Burgess1 1Department of Molecular and Cellular Biology, University of California, Davis, 2Integrative Genetics and Genomics Graduate Group, University of California, Davis Nuclear surface spreads are an indispensable tool for studying chromosome events during meiosis. Here we demonstrate a method to prepare and visualize meiotic chromosomes during prophase I from zebrafish spermatocytes. Neuroscience Reliable Isolation of Central Nervous System Microvessels Across Five Vertebrate Groups Yinyu Yuan*1, Jacquelyn R. Dayton*1, Marie-Lena Freese2, Bryce G. Dorflinger1, Lillian Cruz-Orengo1 1Department of Anatomy, Physiology and Cell Biology, University of California Davis, 2University of Veterinary Medicine Hannover Foundation The goal of this protocol is to isolate microvessels from multiple regions of the central nervous system of lissencephalic and gyrencephalic vertebrates. Bioengineering Prospecting Microbial Strains for Bioremediation and Probiotics Development for Metaorganism Research and Preservation Helena D. M. Villela1, Caren L. S. Vilela1, Juliana M. Assis1, Natascha Varona2, Camille Burke2, David A. Coil2, Jonathan A. Eisen2, Raquel S. Peixoto1,2,3 1LEMM, Laboratory of Molecular Microbial Ecology, Institute of Microbiology Paulo de Góes, Federal University of Rio de Janeiro (UFRJ), 2Genome Center, University of California, Davis, 3IMAM-AquaRio - Rio de Janeiro Aquarium Research Center Pollution affects all biomes. Marine environments have been particularly impacted, especially coral reefs, one of the most sensitive ecosystems on Earth. Bioremediation is the capacity of organisms to degrade contaminants. Here, we describe methodologies to isolate and test microbes presenting bioremediation ability and potential probiotic characteristics for corals. Biochemistry A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products Katherine M. Murphy1, Siwon Chung1, Shruti Fogla1, Hana B. Minsky1, Karen Yong Zhu1, Philipp Zerbe1 1Department of Plant Biology, University of California Davis Here we present easy to use protocols for producing and purifying diterpenoid metabolites through the combinatorial expression of biosynthetic enzymes in Escherichia coli or Nicotiana benthamiana, followed by chromatographic product purification. The resulting metabolites are suitable for various studies including molecular structure characterization, enzyme functional studies, and bioactivity assays. Environment PARbars: Cheap, Easy to Build Ceptometers for Continuous Measurement of Light Interception in Plant Canopies William T. Salter1, Andrew M. Merchant1, Matthew E. Gilbert2, Thomas N. Buckley2 1School of Life and Environmental Sciences, Sydney Institute of Agriculture, University of Sydney, 2Department of Plant Sciences, University of California, Davis Here, we present detailed instructions on how to build and calibrate research quality ceptometers (light sensors that integrate light intensity across many sensors arrayed linearly along a horizontal bar). Immunology and Infection A Mouse Model to Assess Innate Immune Response to Staphylococcus aureus Infection Leif S. Anderson1, Mack B. Reynolds1, Kathryn R. Rivara1, Lloyd S. Miller2, Scott I. Simon1 1Department of Biomedical Engineering, University of California Davis, 2Department of Dermatology, Johns Hopkins University School of Medicine An approach is described for real-time detection of the innate immune response to cutaneous wounding and Staphylococcus aureus infection of mice. By comparing LysM-EGFP mice (which possess fluorescent neutrophils) with a LysM-EGFP crossbred immunodeficient mouse strain, we advance our understanding of infection and the development of approaches to combat infection. Environment Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions Andrew J. Margenot1, Sanjai J. Parikh2, Francisco J. Calderón3 1Department of Crop Sciences, University of Illinois Urbana-Champaign, 2Department of Land, Air and Water Resources, University of California Davis, 3Central Great Plains Resources Management Research, USDA ARS SOM underlies many soil functions and processes, but its characterization by FTIR spectroscopy is often challenged by mineral interferences. The described method can increase the utility of SOM analysis by FTIR spectroscopy by subtracting mineral interferences in soil spectra using empirically obtained mineral reference spectra. Biochemistry Measuring Liver Mitochondrial Oxygen Consumption and Proton Leak Kinetics to Estimate Mitochondrial Respiration in Holstein Dairy Cattle Heidi A. Rossow1, Gabriela Acetoze2, John Champagne1, Jon J. Ramsey3 1Veterinary Medicine Teaching and Research Center, University of California Davis, 2Archer Daniels Midland, 3Department of Molecular Biosciences, University of California Davis Here, we share methods for measuring mitochondrial oxygen consumption, a defining concept of nutritional energetics, and proton leak, the primary cause of inefficiency in mitochondrial generation of ATP. These results can account for 30% of the energy lost in nutrient utilization to help evaluate mitochondrial function. Environment An Optimized Rhizobox Protocol to Visualize Root Growth and Responsiveness to Localized Nutrients Jennifer E. Schmidt1, Carolyn Lowry2, Amelie C.M. Gaudin1 1Department of Plant Sciences, University of California at Davis, 2Department of Natural Resources and the Environment, University of New Hampshire Visualizing and measuring root growth in situ is extremely challenging. We present a customizable rhizobox method to track root development and proliferation over time in response to nutrient enrichment. This method is used to analyze maize genotypic differences in root plasticity in response to an organic nitrogen source. Biochemistry Isolation of Physiologically Active Thylakoids and Their Use in Energy-Dependent Protein Transport Assays Anthony Asher1, Iniyan Ganesan1, Laura Klasek1, Steven M. Theg1 1Department of Plant Biology, University of California - Davis We present protocols herein for high-yield isolation of physiologically active thylakoids and protein transport assays for the chloroplast twin arginine translocation (cpTat), secretory (cpSec1), and signal recognition particle (cpSRP) pathways. Immunology and Infection In Vitro Canine Neutrophil Extracellular Trap Formation: Dynamic and Quantitative Analysis by Fluorescence Microscopy Ronald H.L. Li1, Fern Tablin2 1Department of Veterinary Surgical and Radiological Sciences, School of Veterinary Medicine, University of California, Davis, 2Department of Anatomy, Physiology and Cell Biology, School of Veterinary Medicine, University of California, Davis We describe methods to isolate canine neutrophils from whole blood and visualize NET formation in live neutrophils using fluorescence microscopy. Also described are protocols to quantify NET formation and citrullinated histone H3 (citH3) expression using immunofluorescence microscopy. Medicine An In Vitro Approach to Photodynamic Therapy Evan Austin1,2, Jared Jagdeo1,2,3 1Department of Dermatology, University of California, Davis, 2Dermatology Service, Sacramento VA Medical Center, 3Department of Dermatology, State University of New York, Downstate Medical Center Photodynamic therapy (PDT) is a medical procedure that involves incubation of an exogenously applied photosensitizer (PS) followed by visible light photoactivation to induce apoptosis. We present an in vitro PDT protocol designed to simulate PDT that may be used to study differences in PS incubation and light treatment parameters. Medicine The Perinatal Asphyxiated Lamb Model: A Model for Newborn Resuscitation Payam Vali*1, Sylvia Gugino*2, Carmon Koenigsknecht2, Justin Helman2, Praveen Chandrasekharan2, Munmun Rawat2, Satyan Lakshminrusimha1, Jayasree Nair2 1Department of Pediatrics, University of California Davis, 2Department of Pediatrics, University at Buffalo Invasive instrumentation of the fetal lamb provides accurate physiologic measurements of the transitioning circulation in a model that closely mimics the newly born infant. Bioengineering Surface Functionalization of Hepatitis E Virus Nanoparticles Using Chemical Conjugation Methods Chun Chieh Chen*1, Marie Stark*1, Mo Baikoghli1, R. Holland Cheng1 1Department of Molecular and Cellular Biology, University of California Davis We have engineered the capsid protein of hepatitis E virus as a theranostic nanoparticle (HEVNP). HEVNP self-assembles into a stable icosahedral cage in mucosal delivery. Here, we describe the modification of HEVNPs for tumor targeting by mutating surface-exposed residues to cysteines, which conjugate synthetic ligands that specifically bind tumor cells. Biology In Vitro SUMOylation Assay to Study SUMO E3 Ligase Activity Wan-Shan Yang1, Mel Campbell2, Hsing-Jien Kung2,3,4,5, Pei-Ching Chang1,6 1Institute of Microbiology and Immunology, National Yang-Ming University, 2UC Davis Cancer Center, University of California, Davis, 3Department of Biochemistry and Molecular Medicine, University of California, Davis, 4Institute for Translational Medicine, College of Medical Science and Technology, Taipei Medical University, 5Division of Molecular and Genomic Medicine, National Health Research Institutes, 6Center for Infectious Disease and Cancer Research, Kaohsiung Medical University Unlike ubiquitin ligases, few E3 SUMO ligases have been identified. This modified in vitro SUMOylation protocol is able to identify novel SUMO E3 ligases by an in vitro reconstitution assay. Biology Functional Imaging of Viral Transcription Factories Using 3D Fluorescence Microscopy Christopher P. Chen1, Frank Chuang2, Yoshihiro Izumiya1 1Department of Dermatology, University of California Davis School of Medicine, University of California, Davis, 2Center for Biophotonics, Department of Biochemistry and Molecular Medicine, University of California, Davis Viral transcriptional factories are discrete structures that are enriched with cellular RNA polymerase II to increase viral gene transcription during reactivation. Here, a method to locate sites of actively transcribing viral chromatin in 3D nuclear space by a combination of immunofluorescence staining and in situ RNA hybridization is described. Environment Procedures of Laboratory Fumigation for Pest Control with Nitric Oxide Gas Yong-Biao Liu1, Xiangbing Yang2, Tiffany Masuda1 1Crop Improvement and Protection Unit, USDA-ARS, 2The University of California Davis This paper describes nitric oxide (NO) fumigation protocols for postharvest pest control. Fumigation chambers are flushed with nitrogen (N2) to establish ultralow oxygen conditions before NO is injected. At the end, chambers are flushed with N2 to dilute NO before exposing products to ambient air to prevent exposure to NO2. Behavior Profiling Anti-Neu5Gc IgG in Human Sera with a Sialoglycan Microarray Assay Shani Leviatan Ben-Arye1, Hai Yu2, Xi Chen2, Vered Padler-Karavani1 1Department of Cell Research and Immunology, Tel Aviv University, 2Department of Chemistry, University of California-Davis A sialoglycan microarray assay can be used to evaluate anti-Neu5Gc antibodies in human sera, making it a potential high-throughput diagnostic assay for cancer and other chronic inflammation-mediated human diseases. Bioengineering Treatment of Ligament Constructs with Exercise-conditioned Serum: A Translational Tissue Engineering Model Ann Lee-Barthel1, Keith Baar1,2, Daniel W. D. West1,3 1Department of Neurobiology, Physiology, and Behavior, University of California, Davis, 2Department of Physiology and Membrane Biology, University of California, Davis, 3Faculty of Kinesiology and Physical Education, University of Toronto We present a model of ligament tissue in which three-dimensional constructs are treated with the human exercise-conditioned serum and analyzed for collagen content, function, and cellular biochemistry. Environment Vegetated Treatment Systems for Removing Contaminants Associated with Surface Water Toxicity in Agriculture and Urban Runoff Brian S. Anderson1,2, Bryn M. Phillips1,2, Jennifer P. Voorhees1,2, Michael Cahn3 1Department of Environmental Toxicology, University of California, Davis, 2Marine Pollution Studies Laboratory - Granite Canyon, 3University of California, Cooperative Extension, Monterey County This article summarizes the design attributes and the effectiveness of treatment systems that treat urban stormwater and agriculture irrigation runoff to remove pesticides and other contaminants associated with aquatic toxicity. Engineering Spark Plasma Sintering Apparatus Used for the Formation of Strontium Titanate Bicrystals Lauren A. Hughes1, Klaus van Benthem1 1Department of Materials Science and Engineering, University of California, Davis A viable technique for the formation of strontium titanate bicrystals at high pressure and fast heating rate via the spark plasma sintering apparatus is developed. Biochemistry A Facile and Efficient Approach for the Production of Reversible Disulfide Cross-linked Micelles Yuanpei Li1,2, Gaurav Bharadwaj1, Joyce S. Lee3 1Department of Biochemistry & Molecular Medicine, University of California, Davis, 2UC Davis Comprehensive Cancer Center, University of California, Davis, 3Department of Internal Medicine, Division of Hematology and Oncology, University of California, Davis To deliver cancer drugs to tumor sites with high specificity and reduced side effects, new methods based on nanoparticles are required. Here, we describe disulfide cross-linked micelles that can be easily prepared by hydrogen peroxide-mediated oxidation and are able to dissociate efficiently under a reducing tumor environment to release payloads. Genetics The Use of Induced Somatic Sector Analysis (ISSA) for Studying Genes and Promoters Involved in Wood Formation and Secondary Stem Development Antanas Spokevicius1, Lynette Taylor1, Emma Melder1, Kim Van Beveren1, Josquin Tibbits2, Nicky Creux3,4, Gerd Bossinger1 1School of Ecosystem and Forest Sciences, Faculty of Science, The University of Melbourne, 2Victorian AgriBiosciences Centre, La Trobe University R&D Park, 3College of Biological Sciences, Department of Plant Biology, University of California, Davis, 4Department of Genetics, Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria Here we present a protocol that facilitates the medium to high throughput functional characterization of gene and promoter constructs in tree secondary stem tissue within comparatively short time frames. It is efficient, easy to use and widely applicable to a range of tree species. Genetics Laser-assisted Cytoplasmic Microinjection in Livestock Zygotes Yanina S. Bogliotti1, Marcela Vilarino1, Pablo J. Ross1 1Department of Animal Science, University of California, Davis This protocol shows how to perform cytoplasmic microinjection in farm animal zygotes. This technique can be used to deliver any solution into the one-cell embryo such as genome editing tools to generate knockout animals. Immunology and Infection Visualization of Twitching Motility and Characterization of the Role of the PilG in Xylella fastidiosa Xiangyang Shi1, Hong Lin2 1Department of Plant Science, University of California, Davis, 2Agricultural Research Service, San Joaquin Valley Agricultural Sciences Center, United States Department of Agriculture In this study, a nano-microfluidic flow chamber was employed to visualize and functionally characterize the twitching motility of Xylella fastidiosa, a bacterium that causes Pierce's disease in grapevine. Biology Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk1 Demet Candas1, Lili Qin1, Ming Fan1, Jian-Jian Li1 1Radiation Oncology, University of California, Davis Here, we outline how to study mitochondrial localization of a (cell cycle) kinase, and how to determine its sub-mitochondrial location as well as potential mitochondrial substrates/targets. Forced expression of proteins into the mitochondria provides a useful tool for studying the functional consequences of mitochondrial localization of a protein of interest. Medicine Simultaneous PET/MRI Imaging During Mouse Cerebral Hypoxia-ischemia Yu Ouyang1, Martin S. Judenhofer1, Jeffrey H. Walton1,2, Jan Marik3, Simon P. Williams3, Simon R. Cherry1,4 1Department of Biomedical Engineering, University of California, Davis, 2Nuclear Magnetic Resonance Facility, University of California, Davis, 3Biomedical Imaging, Genentech, Inc, 4Department of Radiology, University of California, Davis The method presented here uses simultaneous positron emission tomography and magnetic resonance imaging. In the cerebral hypoxia-ischemia model, dynamic changes in diffusion and glucose metabolism occur during and after injury. The evolving and irreproducible damage in this model necessitates simultaneous acquisition if meaningful multi-modal imaging data are to be acquired. Biology Microperfusion Technique to Investigate Regulation of Microvessel Permeability in Rat Mesentery Fitz-Roy E. Curry1, Joyce F. Clark1, Roger H. Adamson1 1Department of Physiology & Membrane Biology, University of California Davis The modified Landis technique enables paired measurement of the hydraulic conductivity of individual microvessels in the mesentery of normal and genetically modified rats under control and test conditions using microperfusion techniques. It provides a convenient method to evaluate mechanisms that regulate microvessel permeability and transvascular exchange under physiological conditions. Biology Optical Mapping of Intra-Sarcoplasmic Reticulum Ca2+ and Transmembrane Potential in the Langendorff-perfused Rabbit Heart Lianguo Wang1, Nicole M. De Jesus2, Crystal M. Ripplinger1 1Department of Pharmacology, University of California Davis, 2Departments of Pharmacology and Biomedical Engineering, University of California Davis This article describes the detailed protocol and equipment necessary for dual optical mapping of transmembrane potential (Vm) and free intra-sarcoplasmic reticulum (SR) Ca2+ in the Langendorff-perfused rabbit heart. This method allows for direct observation and quantification of Vm and SR Ca2+ dynamics in the intact heart. Biology Measurement of Extracellular Ion Fluxes Using the Ion-selective Self-referencing Microelectrode Technique Guillaume Luxardi1, Brian Reid1, Fernando Ferreira1,2, Pauline Maillard3, Min Zhao1,4 1Department of Dermatology, Institute for Regenerative Cures, University of California, Davis, 2Departamento de Biologia, Centro de Biologia Molecular e Ambiental, Universidade do Minho, 3Department of Neurology and Center for Neuroscience, University of California, Davis Imaging of Dementia and Aging Laboratory, 4Department of Ophthalmology, Institute for Regenerative Cures, University of California, Davis Transporters in cell membranes allow differential segregation of ions across cell membranes or cell layers and play crucial roles during tissue physiology, repair and pathology. We describe the ion-selective self-referencing microelectrode that allows the measurement of specific ion fluxes at single cells and tissues in vivo. Medicine Calibrated Forceps Model of Spinal Cord Compression Injury Ashley McDonough*1,2, Angela Monterrubio*1,2, Jeanelle Ariza1,2, Verónica Martínez-Cerdeño1,2 1Department of Pathology and Laboratory Medicine, University of California, Davis, 2Institute for Pediatric Regenerative Medicine, Shriners Hospitals for Children (Northern California) Spinal cord injury models should be highly reproducible. We demonstrate that the calibrated forceps compression model of spinal cord injury is an easy to use surgical method for generating reproducible injuries to the murine spinal cord. Biology A Multi-detection Assay for Malaria Transmitting Mosquitoes Yoosook Lee1, Allison M. Weakley1, Catelyn C. Nieman1, Julia Malvick2, Gregory C. Lanzaro1 1Department of Pathology, Microbiology and Immunology, School of Veterinary Medicine, University of California - Davis, 2Veterinary Genetics Laboratory, School of Veterinary Medicine, University of California, Davis Malaria transmitting mosquitoes have a number of epidemiologically important characteristics that can only be detected using molecular techniques. Utilizing a MALDI-TOF based SNP genotyping platform, we developed an assay for simultaneously detecting multiple key traits (species, insecticide resistance, parasite infection and host choice) of malaria vectors. Medicine Focus Formation: A Cell-based Assay to Determine the Oncogenic Potential of a Gene Angel Alvarez1, Gustavo A. Barisone1, Elva Diaz1 1Department of Pharmacology, University of California, Davis The Focus Formation Assay provides a straightforward method to assess the transforming potential of a candidate oncogene. Engineering A Testing Platform for Durability Studies of Polymers and Fiber-reinforced Polymer Composites under Concurrent Hygrothermo-mechanical Stimuli Antonio Gomez1, Robert Pires1, Alyssa Yambao1, Valeria La Saponara1 1Department of Mechanical and Aerospace Engineering, University of California, Davis The durability of polymers and fiber-reinforced polymer composites in service is a critical aspect for their designs and condition-based maintenance. We present a novel low-cost laboratory testing platform for the investigation of the influence of concurrent mechanical and environmental loadings, and may help design more efficient yet safer composite structures. Biology Utilizing Custom-designed Galvanotaxis Chambers to Study Directional Migration of Prostate Cells Hsin-ya Yang1, Thi Dinh La1, R. Rivkah Isseroff1 1Department of Dermatology, Scool of Medicine, University of California, Davis We present a method to apply a physiological electric field to migrating, immortalized prostate cells in a custom-made galvanotaxis chamber. Using this method, we demonstrate that 2 lines of non-tumorigenic prostate cells demonstrate different degrees of migration directionality in the field. Engineering Real-Time DC-dynamic Biasing Method for Switching Time Improvement in Severely Underdamped Fringing-field Electrostatic MEMS Actuators Joshua Small1, Adam Fruehling2, Anurag Garg3, Xiaoguang Liu1, Dimitrios Peroulis3 1Department of Electrical and Computer Engineering, University of California, Davis, 2Digital Light Projection (DLP) Technology Development, Texas Instruments, 3Birck Nanotechnology Center and the Department of Electrical and Computer Engineering, Purdue University The robust device design of fringing-field electrostatic MEMS actuators results in inherently low squeeze-film damping conditions and long settling times when performing switching operations using conventional step biasing. Real-time switching time improvement with DC-dynamic waveforms reduces the settling time of fringing-field MEMS actuators when transitioning between up-to-down and down-to-up states. Immunology and Infection High-throughput Assay to Phenotype Salmonella enterica Typhimurium Association, Invasion, and Replication in Macrophages Jing Wu1, Roberta Pugh1, Richard C. Laughlin1, Helene Andrews-Polymenis2, Michael McClelland3, Andreas J. Bäumler4, L. Garry Adams1 1Department of Veterinary Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, 2Department of Microbial and Molecular Pathogenesis, College of Medicine, Texas A&M University System Health Science Center, 3University of California, Irvine, 4Department of Medical Microbiology & Immunology, School of Medicine, University of California, Davis A high-throughput assay to in vitro phenotype Salmonella or other bacterial association, invasion, and replication in phagocytic cells with high-throughput capacity was developed. The method was employed to evaluate Salmonella gene knockout mutant strains for their involvements in host-pathogen interactions. Environment Colorimetric Paper-based Detection of Escherichia coli, Salmonella spp., and Listeria monocytogenes from Large Volumes of Agricultural Water Bledar Bisha1, Jaclyn A. Adkins2, Jana C. Jokerst3, Jeffrey C. Chandler1, Alma Pérez-Méndez4, Shannon M. Coleman4, Adrian O. Sbodio5, Trevor V. Suslow5, Michelle D. Danyluk6, Charles S. Henry2, Lawrence D. Goodridge7 1Department of Animal Science, University of Wyoming, 2Department of Chemistry, Colorado State University, 3Department of Environmental and Radiological Health Sciences, Colorado State University, 4Department of Animal Sciences, Colorado State University, 5Department of Plant Sciences, University of California, Davis, 6Department of Food Science and Human Nutrition, Citrus Research and Education Center, University of Florida, 7Department of Food Science and Agricultural Chemistry, McGill University A protocol involving integrated concentration, enrichment, and end-point colorimetric detection of foodborne pathogens in large volumes of agricultural water is presented here. Water is filtered through Modified Moore Swabs (MMS), enriched with selective or non-selective media, and detection is performed using paper-based analytical devices (µPAD) imbedded with bacterial-indicative colorimetric substrates. Medicine A Novel in vivo Gene Transfer Technique and in vitro Cell Based Assays for the Study of Bone Loss in Musculoskeletal Disorders Dennis J. Wu1, Neha Dixit1, Erika Suzuki1, Thanh Nguyen2, Hyun Seock Shin1, Jack Davis2, Emanual Maverakis3, Iannis E. Adamopoulos1,2 1Division of Rheumatology, Allergy and Clinical Immunology, University of California, Davis, 2Institute for Pediatric Regenerative Medicine, Shriners Hospitals for Children - Northern California, 3Department of Dermatology, University of California, Davis Differentiation of precursor cells into osteoclasts is regulated by cytokines and growth factors. Here, a novel gene transfer technique for differentiation of osteoclasts in vivo and cell culture protocols for differentiating precursor cells into osteoclasts in vitro as a method to study the effects of cytokines on osteoclastogenesis are described. Environment Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data Andrew J. McElrone1,2, Thomas M. Shapland2, Arturo Calderon2,3, Li Fitzmaurice4,5, Kyaw Tha Paw U4, Richard L. Snyder4 1Crops Pathology and Genetics Research Unit, United States Department of Agriculture-Agricultural Research Service, 2Department of Viticulture & Enology, University of California, Davis, 3Faculty of Agricultural Sciences, University of Chile, 4Atmospheric Science, University of California, Davis, 5URS Corporation Australia Pty. Ltd. Surface renewal is a micrometeorological method that is being used increasingly to determine energy fluxes, but its technical complexity makes it inaccessible to a broad audience. We describe the steps needed to set up and calibrate a surface renewal field station, to acquire and process data, and to correctly interpret results. Neuroscience Electrophysiological Recording in the Brain of Intact Adult Zebrafish Lindsey Johnston1, Rebecca E. Ball2, Seth Acuff3, John Gaudet4, Andrew Sornborger5, James D. Lauderdale2 1Department of Genetics, University of Georgia, 2Department of Cellular Biology, University of Georgia, 3College of Engineering, Architecture and Technology, Oklahoma State University, 4University of Georgia, 5Department of Mathematics, University of California, Davis This paper describes how an adult zebrafish can be immobilized, intubated, and used for in vivo electrophysiological experiments to allow recordings and manipulation of neural activity in an intact animal. Bioengineering Microfabrication of Nanoporous Gold Patterns for Cell-material Interaction Studies Pallavi Daggumati1, Ozge Kurtulus2, Christopher Abbott Reece Chapman3, Damla Dimlioglu1, Erkin Seker1 1Department of Electrical and Computer Engineering, University of California, Davis, 2Department of Chemical Engineering and Materials Science, University of California, Davis, 3Department of Biomedical Engineering, University of California, Davis We report on techniques to micropattern nanoporous gold thin films via stencil printing and photolithography, as well as methods to culture cells on the microfabricated patterns. In addition, we describe image analysis methods to characterize morphology of the material and the cultured cells using scanning electron and fluorescence microscopy techniques. Biology Quantitative Analysis of Autophagy using Advanced 3D Fluorescence Microscopy Chun A. Changou1,2, Deanna L. Wolfson2, Balpreet Singh Ahluwalia2,3, Richard J. Bold4,5, Hsing-Jien Kung5,6, Frank Y.S. Chuang1,2,5 1Department of Biochemistry and Molecular Medicine, University of California, Davis, 2NSF Center for Biophotonics Science & Technology, University of California, Davis, 3University of Tromsø, 4Department of Surgery (Division of Surgical Oncology), University of California, Davis, 5UC Davis Comprehensive Cancer Center, University of California, Davis, 6Department of Biological Chemistry, University of California, Davis Autophagy is a ubiquitous process that enables cells to degrade and recycle proteins and organelles. We apply advanced fluorescence microscopy to visualize and quantify the small, but essential, physical changes associated with the induction of autophagy, including the formation and distribution of autophagosomes and lysosomes, and their fusion into autolysosomes. Biology Using High Resolution Computed Tomography to Visualize the Three Dimensional Structure and Function of Plant Vasculature Andrew J. McElrone1,2, Brendan Choat3, Dilworth Y. Parkinson4, Alastair A. MacDowell4, Craig R. Brodersen5 1U.S. Department of Agriculture, 2Department of Viticulture and Enology, University of California - Davis, 3Hawkesbury Institute for the Environment, University of Western Sydney, 4Advanced Light Source, Lawrence Berkeley National Lab, 5Citrus Research & Education Center, University of Florida High resolution x-ray computed tomography (HRCT) is a non-destructive diagnostic imaging technique that can be used to study the structure and function of plant vasculature in 3D. We demonstrate how HRCT facilitates exploration of xylem networks across a wide range of plant tissues and species. Biology LeafJ: An ImageJ Plugin for Semi-automated Leaf Shape Measurement Julin N. Maloof*1, Kazunari Nozue*1, Maxwell R. Mumbach1, Christine M. Palmer1 1Department of Plant Biology, University of California Davis Demonstration of key methods for high throughput leaf measurements. These methods can be used to accelerate leaf phenotyping when studying many plant mutants or otherwise screening plants by leaf phenotype. Bioengineering On-Chip Endothelial Inflammatory Phenotyping J. Sherrod DeVerse*1, Keith A. Bailey*1, Greg A. Foster1, Vaishali Mittal1, Stuart M. Altman1, Scott I. Simon1, Anthony G. Passerini1 1Department of Biomedical Engineering, University of California, Davis Microfluidic flow chambers etched by photolithography and fabricated from PDMS are applied to probe functional outcomes associated with EC dysfunction and inflammation. In a representative experiment, the ability of differential shear stress to modulate monocytic cell adhesion to cytokine activated EC monolayers is demonstrated. Bioengineering Microfluidic Mixers for Studying Protein Folding Steven A. Waldauer1, Ling Wu1, Shuhuai Yao2, Olgica Bakajin3, Lisa J. Lapidus1 1Department of Physics and Astronomy, Michigan State University, 2Department of Mechanical Engineering, Hong Kong University of Science and Technology, 3Center for Biophotonics, University of California, Davis In this work we explain the fabrication and use of a microfluidic mixer capable of mixing two solutions in ~8 μs. We also demonstrate the use of these mixers with spectroscopic detection using UV fluorescence and fluorescence resonance energy transfer (FRET). Biology Fruit Volatile Analysis Using an Electronic Nose Simona Vallone1, Nathan W. Lloyd2, Susan E. Ebeler3, Florence Zakharov1 1Department of Plant Sciences, University of California, Davis, 2Department of Chemical Engineering and Material Science, University of California, Davis, 3Department of Viticulture and Enology, University of California, Davis A rapid method for volatile compound analysis in fruit is described. The volatile compounds present in the headspace of a homogenate of the sample are rapidly separated and detected with ultra-fast gas chromatography (GC) coupled with a surface acoustic wave (SAW) sensor. A procedure for data handling and analysis is also discussed. Neuroscience Inducing Dendritic Growth in Cultured Sympathetic Neurons Atefeh Ghogha1, Donald A. Bruun1, Pamela J. Lein1 1Department of Molecular Biosciences, University of California, Davis We describe a protocol for using bone morphogenetic protein-7 (BMP-7) or Matrigel to selectively induce dendritic growth in primary sympathetic neurons dissociated from the superior cervical ganglia (SCG) of perinatal rats. Bioengineering Electric Field-controlled Directed Migration of Neural Progenitor Cells in 2D and 3D Environments Xiaoting Meng*1, Wenfei Li*2,3, Fraser Young1, Runchi Gao3, Laura Chalmers3, Min Zhao3, Bing Song1 1School of Dentistry, Cardiff Institute of Tissue Engineering & Repair, Cardiff University, 2Shandong Qianfoshan Hospital, Shandong University School of Medicine, 3Dermatology and Ophthalmology Research, Institute for Regenerative Cures, University of California at Davis This protocol demonstrates methods used to establish 2D and 3D environments in custom-designed electrotactic chambers, which can track cells in vivo/ex vivo using time-lapse recording at the single cell level, in order to investigate galvanotaxis/electrotaxis and other cellular responses to direct current (DC) electric fields (EFs). Biology Quantifying Mixing using Magnetic Resonance Imaging Emilio J. Tozzi1, Kathryn L. McCarthy1, Lori A. Bacca2, William H. Hartt2, Michael J. McCarthy1 1Dept. Food Science and Technology, University of California, Davis, 2Corporate Engineering and Technology Laboratory, Procter & Gamble Company Magnetic resonance imaging (MRI) provides a powerful tool to evaluate the effectiveness of process equipment during operation. We discuss the use of MRI to visualize mixing in a static mixer. The application is relevant to personal care products, but can be applied to a broad range of food, chemical, biomass and biological fluids. Bioengineering Live-cell Imaging of Migrating Cells Expressing Fluorescently-tagged Proteins in a Three-dimensional Matrix Wenting Shih1, Soichiro Yamada1 1University of California, Davis Cellular processes such as cell migration have traditionally been studied on two-dimensional, stiff plastic surfaces. This report describes a technique for directly visualizing protein localization and analyzing protein dynamics in cells migrating in a more physiologically relevant, three-dimensional matrix. Immunology and Infection Rapid Diagnosis of Avian Influenza Virus in Wild Birds: Use of a Portable rRT-PCR and Freeze-dried Reagents in the Field John Y. Takekawa1, Nichola J. Hill1,2, Annie K. Schultz1, Samuel A. Iverson1, Carol J. Cardona3,4, Walter M. Boyce2, Joseph P. Dudley5 1USGS Western Ecological Research Center, 2Wildlife Health Center, University of California, Davis, 3Department of Population Health and Reproduction, University of California, Davis, 4Department of Veterinary and Biomedical Sciences, University of Minnesota, 5Science Applications International Corporation This study describes diagnosis of avian influenza in wild birds using a portable rRT-PCR system. The method takes advantage of freeze-dried reagents to screen wild birds in a non-laboratory setting, typical of an outbreak scenario. Use of molecular tools provides accurate and sensitive alternatives for rapid diagnosis. Biology Transduction of Human Cells with Polymer-complexed Ecotropic Lentivirus for Enhanced Biosafety Bonnie Barrilleaux1, Paul Knoepfler1 1Department of Cell Biology and Human Anatomy, School of Medicine, University of California, Davis Lentiviruses are a valuable research tool for exploring gene function; however, researchers may wish to avoid production of pantropic lentivirus encoding known or suspected oncogenes. As an alternative, we present a safer protocol for use of ecotropic lentivirus on human cells modified to express the ecotropic receptor mSlc7a1. Bioengineering Low-Cost Cryo-Light Microscopy Stage Fabrication for Correlated Light/Electron Microscopy David B. Carlson1, James E. Evans1 1Department of Molecular and Cellular Biology, University of California Davis We demonstrate the fabrication of a low-cost cryogenic stage designed to fit most reflected light microscopes. This lab-built cryogenic stage enables efficient and reliable correlative imaging between cryo-light and cryo-electron microscopy. Neuroscience Lentivirus-mediated Genetic Manipulation and Visualization of Olfactory Sensory Neurons in vivo Benjamin Sadrian1, Huaiyang Chen1, Qizhi Gong1 1Department of Cell Biology and Human Anatomy, School of Medicine, University of California, Davis We present a lentiviral technique for genetic manipulation and visualization of single olfactory sensory neuron axon and its terminal arborization in vivo. Bioengineering Rejection of Fluorescence Background in Resonance and Spontaneous Raman Microspectroscopy Zachary J. Smith*1, Florian Knorr*1, Cynthia V. Pagba1, Sebastian Wachsmann-Hogiu1,2 1Center for Biophotonics Science and Technology, University of California, Davis, 2Department of Pathology and Laboratory Medicine, University of California, Davis We discuss the construction and operation of a complex nonlinear optical system that uses ultrafast all-optical switching to isolate Raman from fluorescence signals. Using this system we are able to successfully separate Raman and fluorescence signals utilizing pulse energies and average powers that remain biologically safe. Neuroscience Electrophysiological Measurements from a Moth Olfactory System Zainulabeuddin Syed1, Walter S. Leal1 1Department of Entomology, University of California, Davis Insect olfactory systems provide unique opportunities for recording odorant-induced responses in the forms of electroantennograms (EAG) and single sensillum recordings (SSR), which are summed responses from all odorant receptor neurons (ORNs) located on the antenna and from those housed in individual sensilla, respectively. Biology DNA Microarrays: Sample Quality Control, Array Hybridization and Scanning Elva Diaz1, Gustavo A. Barisone1 1Department of Pharmacology, University of California, Davis We demonstrate the use of DNA microarrays for expression profiling of the nervous system. We describe RNA quality control, sample labeling, and array hybridization and scanning. Bioengineering Measurement of Bioelectric Current with a Vibrating Probe Brian Reid1, Min Zhao1 1Dermatology, University of California, Davis The manufacture, calibration and use of non-invasive vibrating probes to measure bioelectric current in various biological systems is described. Immunology and Infection Transfection and Mutagenesis of Target Genes in Mosquito Cells by Locked Nucleic Acid-modified Oligonucleotides Nazzy Pakpour1, Kong Wai Cheung1, Lattha Souvannaseng1, Jean-Paul Concordet2, Shirley Luckhart1 1Department of Medical Microbiology and Immunology, University of California, Davis, 2Département Génétique et Développement, Institut Cochin, Université Paris Descartes Oligonucleotides can be used to site specifically substitute a single nucleotide of transfected target genes in both Anopheles gambiae and Anopheles stephensi cells. Biology Whole-Cell Recording of Calcium Release-Activated Calcium (CRAC) Currents in Human T Lymphocytes Pratima Thakur1, Alla F. Fomina1 1Department of Physiology and Membrance Biology, University of California, Davis We provide a step-by-step protocol for whole-cell patch clamp recording of Calcium Release-Activated Calcium (CRAC) currents in peripheral blood mononuclear cell-derived human T lymphocytes. Biology Differential Imaging of Biological Structures with Doubly-resonant Coherent Anti-stokes Raman Scattering (CARS) Tyler J. Weeks1, Thomas R. Huser1,2 1Center for Biophotonics, University of California, Davis, 2Department of Internal Medicine, University of California, Davis A combination of three single wavelength short-pulsed lasers is used to generate coherent anti-Stokes Raman scattering (CARS) and doubly-resonant CARS (DR-CARS). The difference between these signals provides enhanced sensitivity for otherwise difficult to detect coherent Raman signals, enabling imaging of weak Raman scatterers. Immunology and Infection Visualizing Cell-to-cell Transfer of HIV using Fluorescent Clones of HIV and Live Confocal Microscopy Benjamin Dale1, Gregory P. McNerney2, Deanna L. Thompson2, Wolfgang Hübner3, Thomas Huser2, Benjamin K. Chen1 1Division of Infectious Diseases, Department of Medicine, Immunology Institute, Mount Sinai School of Medicine, 2NSF Center for Biophotonics, University of California, Davis, 3Structural and Computational Biology Unit, European Molecular Biology Laboratory This visualized experiment is a guide for utilizing a fluorescent molecular clone of HIV for live confocal imaging experiments. Neuroscience Assaying Locomotor Activity to Study Circadian Rhythms and Sleep Parameters in Drosophila Joanna C. Chiu1,2, Kwang Huei Low1,3, Douglas H. Pike1, Evrim Yildirim1,3, Isaac Edery1,3 1Center for Advanced Biotechnology and Medicine, Rutgers University, 2Current Address: Department of Entomology, College of Agricultural and Environmental Sciences, University of California, Davis, 3Department of Molecular Biology and Biochemistry, Rutgers University We describe procedures for recording daily locomotor activity rhythms of Drosophila and subsequent data analysis. Locomotor activity rhythms are a reliable behavioral output of animal circadian clocks and are used as the standard readout of clock function when studying circadian mutants or examining how the environment regulates the circadian system. Biology Differentiation of Embryonic Stem Cells into Oligodendrocyte Precursors Peng Jiang1, Vimal Selvaraj1, Wenbin Deng1 1Department of Cell Biology and Human Anatomy Institute for Pediatric Regenerative Medicine, School of Medicine, University of California, Davis We describe a small molecule-based protocol for differentiation of mouse embryonic stem cells into oligodendrocyte precursor cells (OPCs). This protocol generates Olig2+NG2+ OPCs with high efficiency by 30 days of differentiation. We also describe a method to generate "spiking" OPCs that can fire action potentials. Neuroscience Mouse Models of Periventricular Leukomalacia Yan Shen1, Jennifer M. Plane1, Wenbin Deng1 1Department of Cell Biology and Human Anatomy Institute for Pediatric Regenerative Medicine School of, University of California, Davis We established mouse models of periventricular leukomalacia (PVL), the predominant brain injury in premature infants characterized by periventricular white matter lesions. Hypoxia/ischemia with/without systemic infection are the primary causes of PVL. Unilateral carotid ligation and hypoxia exposure with/without lipopolysaccharide injection creates PVL-like lesions in P6 mice. Biology Microinjection Techniques for Studying Mitosis in the Drosophila melanogaster Syncytial Embryo Ingrid Brust-Mascher1, Jonathan M. Scholey1 1Department of Molecular and Cellular Biology, University of California, Davis This protocol describes the use of microinjection and high resolution imaging in the Drosophila melanogaster syncytial embryo to study mitosis. Biology A High-Throughput Method For Zebrafish Sperm Cryopreservation and In Vitro Fertilization Bruce W. Draper1, Cecilia B. Moens2 1Molecular and Cellular Biology, University of California, Davis, 2Division of Basic Science, Fred Hutchinson Cancer Research Center - FHCRC This is a high-throughput sperm cryopreservation protocol for zebrafish. Sperm cryopreserved using this protocol has an average of 25% fertility in subsequent vitro fertilization and is stable over many years. Biology Blood Collection from the American Horseshoe Crab, Limulus Polyphemus Peter Armstrong1,2, Mara Conrad3 1Department of Molecular and Cell Biology, University of California, Davis, 2Marine Biological Laboratory - MBL- woods hole, 3Department of Biological Sciences, Hunter College of CUNY The American horseshoe crab, Limulus polyphemus, is arguably the most convenient source for large quantities of blood of any invertebrate. The blood is simple in composition, with only one cell-type in the general circulation, the granular amebocyte, and only three abundant proteins in the plasma, hemocyanin, the C-reactive proteins, and α2-macroglobulin. Blood is collected from the heart and the blood cells and plasma are separated by centrifugation. Biology Preparation of Gene Gun Bullets and Biolistic Transfection of Neurons in Slice Culture Georgia Woods1, Karen Zito1 1Center for Neuroscience, University of California, Davis We describe a method for preparing DNA coated gold bullets and demonstrate the use of such bullets to biolistically transfect neurons in cultured hippocampal slices. Biology Patterning Cells on Optically Transparent Indium Tin Oxide Electrodes Sunny Shah1, Alexander Revzin1 1Department of Biomedical Engineering, University of California, Davis Non-fouling PEG silane monolayer was desorbed from individually addressable ITO electrodes on glass by application of a reductive potential. Electrochemical stripping of PEG-silane layer from ITO microelectrodes allowed for cell adhesion to take place in a spatially defined fashion, with cellular patterns corresponding closely to electrode patterns.
