JoVE General
Assembly, Tuning and Use of an Apertureless Near Field Infrared Microscope for Protein Imaging
1Department of Chemistry, University of Toronto, 2Department of Chemistry, University of Wisconsin, 3Department of Chemistry, Duke University
The assembly of a nearfield infrared microscope for imaging protein aggregates is described.
JoVE Neuroscience
In vivo Laser Axotomy in C. elegans
A protocol to cut neurons in C. elegans with a MicroPoint pulsed laser is presented. We describe setting up the system, immobilizing worms, and severing labeled neurons. Advantages include a relatively low-cost system and the ability to sever neuronal processes or ablate cells in vivo.
JoVE Immunology and Infection
Saliva, Salivary Gland, and Hemolymph Collection from Ixodes scapularis Ticks
1Microbiology and Pathogenesis Activity, Bacterial Diseases Branch, Division of Vector-Borne Diseases, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, 2Tick-Borne Diseases Activity, Bacterial Diseases Branch, Division of Vector-Borne Diseases, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention
The collection of infected tick hemolymph, salivary glands, and saliva is important to study how tick-borne pathogens cause disease. In this protocol we demonstrate how to collect hemolymph and salivary glands from feeding Ixodes scapularis nymphs. We also demonstrate saliva collection from female I. scapularis adults.
JoVE General
Live Cell Imaging of F-actin Dynamics via Fluorescent Speckle Microscopy (FSM)
Department of Cell Biology, Scripps Institute
Selection, microinjection, and imaging of fluorescently-labeled F-actin via fluorescent speckle microscopy (FSM).
JoVE General
Quantitative Analysis of Random Migration of Cells Using Time-lapse Video Microscopy
1Department of Biochemistry and Molecular Biology, LSU School of Medicine, 2Department of Oral Biology, LSU School of Dentistry, 3Stanley S. Scott Cancer Center, LSU School of Medicine
This method allows monitoring of cells in real time and quantitative measurements of different cell migration parameters such as speed, displacement, and velocity. Unlike the traditional methods, this real time approach is not based on endpoint quantitative migration measurements; instead it allows monitoring and calculating different parameters continuously.
JoVE General
A Semi-quantitative Approach to Assess Biofilm Formation Using Wrinkled Colony Development
Department of Microbiology and Immunology, Loyola University Medical Center
We provide a simple, semi-quantitative method to investigate biofilm formation in vitro. This method takes advantage of the Zeiss stemi 2000-C Dissecting Microscope (with camera attachment) to monitor both the timing and pattern of biofilm formation, as assessed by the development of wrinkled colonies.
JoVE General
Optical Scatter Microscopy Based on Two-Dimensional Gabor Filters
Department of Biomedical Engineering, Rutgers University
We demonstrate a dark-field microscopy method based on Gabor-like filtering to measure subcellular dynamics within single living cells. The technique is sensitive to alterations in the structure of organelles, such as mitochondrial fragmentation.
JoVE Clinical and Translational Medicine
Guidelines for Elective Pediatric Fiberoptic Intubation
1Department of Anesthesia, St. Jude Children's Research Hospital, 2Department of Anesthesia, Children's Hospital of Michigan, 3Department of Anesthesiology, Children's Hospital of Michigan
We describe guidelines to perform a safe and efficient elective fiberoptic intubation in pediatric patients while maintaining spontaneous ventilation.
JoVE General
Activation of Apoptosis by Cytoplasmic Microinjection of Cytochrome c
1Department of Cell and Developmental Biology, Neuroscience Center, University of North Carolina, 2Curriculum in Neurobiology, Neuroscience Center, University of North Carolina
In this protocol, we describe the direct cytoplasmic microinjection of cytochrome c protein into fibroblasts and primary sympathetic neurons. This technique allows for the introduction of cytochrome c protein into the cytoplasm of cells and mimics the release of cytochrome c from mitochondria, which occurs during apoptosis.
JoVE General
Major Components of the Light Microscope
Core Optical Imaging Facility, University of Texas Health Science Center at San Antonio (UTHSCSA)
The light microscope is a basic tool for the cell biologist, who should have a thorough understanding of how it works, how it should be aligned for different applications, and how it should be maintained as required to obtain maximum image-forming capacity and resolution. The components of the microscope are described in detail here.
JoVE Neuroscience
Whole Cell Recording from an Organotypic Slice Preparation of Neocortex
Department of Anatomy and Neurobiology, University of Tennessee Health Science Center
This is a protocol to prepare and maintain a neocortical slice preparation in organotypic culture for the purpose of making electrical recordings from pyramidal neurons.
JoVE Bioengineering
Microfluidic Mixers for Studying Protein Folding
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).
JoVE Immunology and Infection
Real-time Imaging of Heterotypic Platelet-neutrophil Interactions on the Activated Endothelium During Vascular Inflammation and Thrombus Formation in Live Mice
1Department of Pharmacology, University of Illinois at Chicago, 2Department of Anesthesiology, University of Illinois at Chicago
Here we report an experimental technique of fluorescence intravital microscopy to visualize heterotypic platelet-neutrophil interactions on the activated endothelium during vascular inflammation and thrombus formation in live mice. This microscopic technology will be valuable to study the molecular mechanism of vascular disease and to test pharmacologic agents under pathophysiological conditions.
JoVE Neuroscience
Odorant-induced Responses Recorded from Olfactory Receptor Neurons using the Suction Pipette Technique
1Monell Chemical Senses Center, 2Department of Physiology, Development & Neuroscience, Physiological Laboratory, University of Cambridge
Olfactory receptor neurons (ORNs) convert odor signals first into a receptor current that in turn triggers action potentials that are conveyed to second order neurons in the olfactory bulb. Here we describe the suction pipette technique to record simultaneously the odorant-induced receptor current and action potentials from mouse ORNs.
JoVE Clinical and Translational Medicine
Mesenteric Artery Contraction and Relaxation Studies Using Automated Wire Myography
1Julius L. Chambers Biomedical/Biotechnology Research Institute, North Carolina Central University, Durham, 2Department of Biology, North Carolina Central University, Durham, 3Department of Physiology & Pharmacology and Hypertension & Vascular Research Center, Wake Forest University School of Medicine
An automated myography method for force measurements in isolated mesenteric arteries is described. It employs a Mulvany-Halpern Auto Dual Wire Myograph 510A to determine responses to phenylephrine and extracellular calcium. The method allows consistent determination of isometric responses to agonists in small vessels of diameters of 60 - 300 μm, independently.
JoVE Clinical and Translational Medicine
High-Resolution Endocardial and Epicardial Optical Mapping in a Sheep Model of Stretch-Induced Atrial Fibrillation
Center for Arrhythmia Research. Internal Medicine, University of Michigan
This report provides a detailed description of the methodology and results of simultaneous endocardial and epicardial optical mapping of electrical excitation in the intact left atrium of a Langendorff-perfused sheep heart during stretch-induced atrial fibrillation.
JoVE Immunology and Infection
Right Ventricular Systolic Pressure Measurements in Combination with Harvest of Lung and Immune Tissue Samples in Mice
1Department of Environmental Medicine, New York University School of Medicine, Tuxedo, 2Division of Allergy, Pulmonary, & Critical Care Medicine, Department of Medicine, Vanderbilt University Medical Center, 3Division of Pulmonary Medicine, New York University School of Medicine
A specific and rapid protocol to simultaneously investigate right heart function, lung inflammation, and the immune response is described as a learning tool. Video and figures describe physiology and microdissection techniques in an organized team-approach that is adaptable to be used for small to large sized studies.
JoVE Neuroscience
A Molecular Readout of Long-term Olfactory Adaptation in C. elegans
1Department of Biological Sciences and Institute for Neuroscience, George Washington University, 2Fred Hutchinson Cancer Research Center, 3Department of Cell and Tissue Biology, University of California San Francisco
Here we describe a molecular readout of long-term olfactory adaptation in Caenorhabditis elegans. The Protein Kinase G, EGL-4, is necessary for stable adaptation responses in the primary sensory neuron pair called AWC. During prolonged odor exposure EGL-4 translocates from the cytosol to nucleus of the AWC.
JoVE General
Microgavage of Zebrafish Larvae
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill
We present a novel method for microgavage of larval zebrafish utilizing standard embryo microinjection and stereomicroscopy equipment. We demonstrate that microgavage is a safe and efficient technique useful for delivering controlled amounts of diverse materials specifically into the larval zebrafish intestinal lumen.
JoVE Bioengineering
Video-rate Scanning Confocal Microscopy and Microendoscopy
1Program in Biophysics, Harvard University, 2Division of Health Sciences and Technology, Harvard-MIT, 3Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School
The complete construction of a custom, real-time confocal scanning imaging system is described. This system, which can be readily used for video-rate microscopy and microendoscopy, allows for an array of imaging geometries and applications not accessible using standard commercial confocal systems, at a fraction of the cost.
JoVE General
Dendra2 Photoswitching through the Mammary Imaging Window
1Department of Anatomy and Structural Biology, Albert Einstein College of Medicine - Yeshiva University, 2Gruss Lipper Biophotonics Center, Albert Einstein College of Medicine - Yeshiva University, 3Hubrecht Institute-KNAW and University Medical Center Utrecht
Intravital photoswitching and tracking of Dendra2-labeled tumor cells through the Mammary Imaging Window is a technique which allows us to image the metastatic behavior of tumor cells in chosen tumor microenvironments over a timescale of days.
JoVE General
A Reverse Genetic Approach to Test Functional Redundancy During Embryogenesis
Department of Surgery, Weill Cornell Medical College of Cornell University
Gene function can be obscured in loss-of-function experiments if there is compensation by another gene. The zebrafish model provides a relatively high-throughput means to reveal such functional redundancy in living embryos.
JoVE General
Computer-assisted Large-scale Visualization and Quantification of Pancreatic Islet Mass, Size Distribution and Architecture
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.
JoVE Bioengineering
Measurement of Bioelectric Current with a Vibrating Probe
Dermatology, University of California, Davis
The manufacture, calibration and use of non-invasive vibrating probes to measure bioelectric current in various biological systems is described.
JoVE Neuroscience
Dissection, Culture, and Analysis of Xenopus laevis Embryonic Retinal Tissue
Department of Biology, College of William and Mary
Xenopus laevis provides an ideal model system for studying cell fate specification and physiological function of individual retinal cells in primary cell culture. Here we present a technique for dissecting retinal tissues and generating primary cell cultures that are imaged for calcium activity and analyzed by in situ hybridization.
JoVE Neuroscience
Transcriptome Analysis of Single Cells
1Department of Pharmacology, University of Pennsylvania, 2The Penn Genome Frontiers Institute, University of Pennsylvania
In this article we describe a simple method for the harvesting of single cells from rat primary neuronal cultures and subsequent transcriptome analysis using aRNA amplification. This approach is generalizable to any cell type.
JoVE Neuroscience
How to Create and Use Binocular Rivalry
1Department of Psychology, New York University, 2Centre for Neural Science, New York University, 3Department of Psychology, Princeton University, 4Neuroscience Institute, Princeton University
Binocular rivalry occurs when the eyes are presented with different images at the same location: one image dominates while the other is suppressed, and dominance alternates periodically. Rivalry is useful for investigating perceptual selection and visual awareness. Here we describe several easy methods for creating and using binocular rivalry stimuli.
JoVE General
Adult and Embryonic Skeletal Muscle Microexplant Culture and Isolation of Skeletal Muscle Stem Cells
School of Biosciences, University of Birmingham
The micro-dissected explants technique is a robust and reliable method for isolating proliferative skeletal muscle cells from juvenile, adult or embryonic muscles as a source of skeletal muscle stem cells. Uniquely, these cells have been clonally derived to produce skeletal muscle stem cell lines used for in vivo transplantation.
JoVE General
Two- and Three-Dimensional Live Cell Imaging of DNA Damage Response Proteins
1Department of Radiation Oncology, Virginia Commonwealth University, 2Department of Biochemistry & Molecular Biology, Virginia Commonwealth University, 3Department of Anatomy & Neurobiology, Virginia Commonwealth University, 4Massey Cancer Center, Virginia Commonwealth University
This protocol describes a method for visualizing a DNA double-strand break signaling protein activated in response to DNA damage as well as its localization during mitosis.
JoVE Neuroscience
Simultaneous Pre- and Post-synaptic Electrophysiological Recording from Xenopus Nerve-muscle Co-cultures
1Department of Physiology, David Geffen School of Medicine at UCLA, 2Natural Science Division, Pepperdine University
This video demonstrates the procedures used to grow primary cultures of embryonic Xenopus nerve and muscle cells and the usefulness of this preparation for making simultaneous pre- and post-synaptic patch clamp recordings.
JoVE General
Monitoring Plasmid Replication in Live Mammalian Cells over Multiple Generations by Fluorescence Microscopy
Department of Oncology, University of Wisconsin - Madison
A method of observing individual DNA molecules in live cells is described. The technique is based on the binding of a fluorescently tagged lac repressor protein to binding sites engineered into the DNA of interest. This method can be adapted to follow many recombinant DNAs in live cells over time.
JoVE Neuroscience
Cerebral Blood Oxygenation Measurement Based on Oxygen-dependent Quenching of Phosphorescence
1Optics Division, Athinoula A. Martinos Center for Biomedical Imaging, Department of Radiology, Massachusetts General Hospital and Harvard Medical School, 2Department of Biochemistry and Biophysics, University of Pennsylvania, 3Neuroprotection Research Laboratory, Departments of Radiology and Neurology, Massachusetts General Hospital and Harvard Medical School, 4Departments of Neurosciences and Radiology, University of California
We present an experimental procedure for measuring the partial pressure of oxygen (pO2) in cerebral vasculature based on oxygen-dependent quenching of phosphorescence. Animal preparation and imaging procedures were outlined for both large field of view CCD-based imaging of pO2 in rats and 2-photon excitation based imaging of pO2 in mice.
JoVE General
Calcium Imaging of Cortical Neurons using Fura-2 AM
1Department of Neurobiology, Stanford University, 2Department of Neurobiology, Stanford University School of Medicine
Calcium signals play a key role in many cellular processes including gene expression, survival and differentiation. Here we demonstrate how to perform calcium imaging using Fura-2 AM. Calcium imaging is a valuable tool to study the regulation of intracellular calcium in real time and its regulation of signaling cascades.
JoVE General
Live Imaging of Cell Extrusion from the Epidermis of Developing Zebrafish
Department of Oncological Sciences, Huntsman Cancer Institute, University of Utah
Dying cells are extruded from epithelial tissues by concerted contraction of neighboring cells without disrupting barrier function. The optical clarity of developing zebrafish provides an excellent system to visualize extrusion in living epithelia. Here we describe methods to induce and image extrusion in the larval zebrafish epidermis at cellular resolution.
JoVE Bioengineering
Low-Cost Cryo-Light Microscopy Stage Fabrication for Correlated Light/Electron Microscopy
Department 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.
JoVE Neuroscience
High-resolution Live Imaging of Cell Behavior in the Developing Neuroepithelium
1Neural Development Group, Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee, UK, 2Wellcome Trust Centre for Gene Regulation and Expression, College of Life Sciences, University of Dundee, Dundee, UK
Imaging embryonic tissue in real-time is challenging over long periods of time. Here we present an assay for monitoring cellular and sub-cellular changes in chick spinal cord for long periods with high spatial and temporal resolution. This technique can be adapted for other regions of the nervous system and developing embryo.
JoVE Applied Physics
Synthesis and Operation of Fluorescent-core Microcavities for Refractometric Sensing
Department of Physics, University of Alberta
Fluorescent-core microcavity sensors employ a high-index quantum-dot coating in the channel of silica microcapillaries. Changes in the refractive index of fluids pumped into the capillary channel cause shifts in the microcavity fluorescence spectrum that can be used to analyze the channel medium.
JoVE Neuroscience
Patch Clamp Recordings in Inner Ear Hair Cells Isolated from Zebrafish
Natural Science Division, Pepperdine University
The purpose of this video is to demonstrate procedures for obtaining healthy, intact hair cells from the inner ear organs of adult zebrafish and then using them for patch clamp studies aimed at characterizing the biophysical properties of their voltage-gated channels.
JoVE Neuroscience
Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster
1Institute of Genetics, University of Mainz, 2Department of Anatomy and Neuroscience, University of Melbourne
We present a technique for labeling single neurons in the central nervous system (CNS) of Drosophila embryos, which allows the analysis of neuronal morphology by either transmitted light or confocal microscopy.
JoVE Clinical and Translational Medicine
Live Imaging of Drug Responses in the Tumor Microenvironment in Mouse Models of Breast Cancer
1Watson School of Biological Sciences, 2Cold Spring Harbor Laboratory, 3Departments of Medical Genetics, University of Oslo and Oslo University Hospital
We describe a method for imaging response to anti-cancer treatment in vivo and at single cell resolution.
JoVE General
The ex vivo Isolated Skeletal Microvessel Preparation for Investigation of Vascular Reactivity
An ex vivo preparation is described for isolation of the largest gracilis muscle resistance arterioles for interrogation of both vascular responses to vasoactive stimuli and the assessment of basic structural properties via passive wall mechanics.
JoVE Neuroscience
Postsynaptic Recordings at Afferent Dendrites Contacting Cochlear Inner Hair Cells: Monitoring Multivesicular Release at a Ribbon Synapse
1Department of Otolaryngology-Head and Neck Surgery., The Johns Hopkins School of Medicine, 2Instituto de Investigaciones en Ingeniería Genética y Biología Molecular, Consejo Nacional de Investigaciones Científicas y Técnicas
Whole-cell patch-clamp recordings from auditory nerve fiber dendrites at the inner hair cell ribbon synapse in the mammalian cochlea.
More Results...
