Chemotaxis:
The movement of cells or organisms toward or away from a substance in response to its concentration gradient.
JoVE Neuroscience
C. elegans Positive Butanone Learning, Short-term, and Long-term Associative Memory Assays
1Department of Molecular Biology, Lewis-Sigler Institute for Integrative Genomics, Princeton University, 2Lewis-Sigler Institute for Integrative Genomics, Princeton University
Here we describe methods to test C. elegans associative learning and short- and long-term associative memory. These population assays employ the worms abilities to chemotax toward volatile odorants, and form positive associations upon pairing food with the chemoattractant butanone. Increasing the number of conditioning periods induces long-term memory.
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 Immunology and Infection
Tracking Neutrophil Intraluminal Crawling, Transendothelial Migration and Chemotaxis in Tissue by Intravital Video Microscopy
Department of Pharmacology, University of Saskatchewan
We describe a protocol of brightfield intravital microscopy for measuring dynamic neutrophil-endothelial cell interactions during neutrophil recruitment in response to the source of a neutrophil chemoattractant in vivo. Neutrophil intraluminal crawling, transendothelial migration and chemotaxis in mouse cremaster muscle tissue are visualized with time-lapsed video photography and tracked with ImageJ.
JoVE General
A Microfluidic Device for Quantifying Bacterial Chemotaxis in Stable Concentration Gradients
1McFerrin Department of Chemical Engineering, Texas A&M University, 2Department of Biology, Texas A&M University, 3Department of Biomedical Engineering, Texas A&M University
This protocol describes the development of a microfluidic device for investigating bacterial chemotaxis in stable concentration gradients of chemoeffectors.
JoVE General
C. elegans Chemotaxis Assay
1Life Sciences, Queen's University, 2Department of Chemical Engineering, Queen's University, 3Department of Biology, Queen's University
A method of quantitatively evaluating the chemotactic response of Caenorhabditis elegans is described. A chemotactic index (CI) was employed as a way to precisely evaluate the response of worms to certain targets, and serve as a platform of comparison between strains and compounds of interest.
JoVE General
Imaging G-protein Coupled Receptor (GPCR)-mediated Signaling Events that Control Chemotaxis of Dictyostelium Discoideum
Here, we describe detailed live cell imaging methods for investigating chemotaxis. We present fluorescence microscopic methods to monitor spatiotemporal dynamics of signaling events in migrating cells. Measurement of signaling events permits us to further understand how a GPCR-signaling network achieves gradient sensing of chemoattractants and controls directional migration of eukaryotic cells.
JoVE General
Two Types of Assays for Detecting Frog Sperm Chemoattraction
1Department of Animal Sciences, University of Illinois, Urbana-Champaign, 2School of Life Sciences, Arizona State University
Eggs and the extracellular coatings around eggs frequently release peptides, proteins and small molecules that communicate with sperm to guide them to the egg thereby promoting fertilization. Using frog sperm we describe and compare two classes of assays used to detect sperm chemoattraction – sperm accumulation assays and sperm tracking assays.
JoVE General
High-resolution Measurement of Odor-Driven Behavior in Drosophila Larvae
Laboratory of Neurogenetics and Behavior, Rockefeller University
In this video article, we describe a new method allowing the construction of odorant gradients with stable and controllable geometries. We briefly illustrate how these gradients can be used to screen for olfactory defects (full and partial anosmia) and to study more subtle features of chemotaxis behavior.
JoVE Neuroscience
A Caenorhabditis elegans Model System for Amylopathy Study
We describe methods to study aspects of amylopathies in the worm C. elegans. We show how to construct worms expressing human Aβ42 in neurons and how to test their function in behavioral assays. We further show how to obtain primary neuronal cultures that can be used for pharmacological testing.
JoVE General
Measurement of Cellular Chemotaxis with ECIS/Taxis
1Molecular and Cell Biology, University of Connecticut, 2University of Connecticut
The ECIS/Taxis system is an automated, real-time assay that measures cellular chemotaxis. In this assay, cells move beneath a layer of agarose to arrive at a target electrode. Cellular movement is measured by the onset of resistance to AC current 0.
JoVE General
A Gradient-generating Microfluidic Device for Cell Biology
We describe a protocol for the microfabrication of the gradient-generating microfluidic device that can generate spatial and temporal gradients in well-defined microenvironment. In this approach, the gradient-generating microfluidic device can be used to study directed cell migration, embryogenesis, wound healing, and cancer metastasis.
JoVE Editorial
September 2011: This Month in JoVE
Here are some highlights from the September 2011 Issue of Journal of Visualized Experiments (JoVE).
JoVE Bioengineering
Creating Adhesive and Soluble Gradients for Imaging Cell Migration with Fluorescence Microscopy
1Centre for Vascular Research and Australian Centre for Nanomedicine, The University of New South Wales, 2School of Chemistry and Australian Centre for Nanomedicine, The University of New South Wales
A method for the assembly of adhesive and soluble gradients in a microscopy chamber for live cell migration studies is described. The engineered environment combines antifouling surfaces and adhesive tracks with solution gradients and therefore allows one to determine the relative importance of guidance cues.
JoVE General
Recording Multicellular Behavior in Myxococcus xanthus Biofilms using Time-lapse Microcinematography
1Department of Environmental Health Sciences, University of South Carolina (USC), 2Department of Biology, Syracuse University
To study Myxococcus xanthus swarm behavior, we have designed a time-lapse microcinematography protocol that can be modified for different assays. It employs standard growth conditions adapted for microscopy, and yields reproducible results by the use of inexpensive, reusable silicone gaskets. We have used this method to quantify multicellular chemotaxis.
JoVE Neuroscience
Analysis of Trunk Neural Crest Cell Migration using a Modified Zigmond Chamber Assay
1Department of Biology, California State University, Northridge, 2Centro de Biología Celular y Molecular, Universidad Nacional de Córdoba
An approach to analyze the migration of explanted cells (trunk neural crest cells) is described. This method is inexpensive, gentle, and capable of distinguishing chemotaxis from both chemokinesis and other influences on migratory polarity such as those derived from cell-cell interactions within the primary trunk neural crest cell culture.
JoVE Immunology and Infection
Real-time Imaging of Leukotriene B4 Mediated Cell Migration and BLT1 Interactions with β-arrestin
Microbiology and Immunology, James Graham Brown Cancer Center, University of Louisville
This paper describes the methodology to determine the chemotactic response of leukocytes to specific ligands and identify interactions between the cell surface receptors and cytosolic proteins using live cell imaging techniques.
JoVE Neuroscience
Lectin-based Isolation and Culture of Mouse Embryonic Motoneurons
1Institute for Cellmorphology and molecular Neurobiology, Group for Cellbiology, Ruhr-University Bochum, 2Institute for Clinical Neurobiology, University of Wuerzburg
An alternative way of isolating mouse embryonic motoneurons from the spinal cord is described. The method takes into account the fact that lectin can bind to the low affinity nerve growth factor receptor p75NTR. This lectin-based preplating allows a purification similar to that with a specific antibody against the p75NTR.
JoVE Editorial
January 2012: This Month in JoVE
Here are some highlights from the January 2012 Issue of Journal of Visualized Experiments (JoVE).
JoVE General
Protein Crystallization for X-ray Crystallography
Molecular Biochemistry and Biophysics, Yale University
The 3-D structure of a molecule provides a unique understanding of how the molecule functions. The principal method for structure determination at near-atomic resolution is X-ray crystallography. Here, we demonstrate the current methods for obtaining three-dimensional crystals of any given macromolecule that are suitable for structure determination by X-ray crystallography.
JoVE Immunology and Infection
Ex vivo Imaging of T Cells in Murine Lymph Node Slices with Widefield and Confocal Microscopes
1Institut Cochin, Université Paris Descartes, CNRS (UMR 8104), 2Inserm, U1016, Paris, France
This protocol describes a method to image fluorescent T cells introduced into lymph node slices. The technique permits real-time analyses of T cell migration with traditional widefield fluorescence or confocal microscopes.
JoVE General
Generation of Human CD40-activated B cells
In this video we present the ex vivo generation and expansion of human CD40-activated B cells (CD40-B) from peripheral blood mononuclear cells (PBMC) by stimulation with CD40 ligand and interleukin-4.
JoVE General
Assessing Neural Stem Cell Motility Using an Agarose Gel-based Microfluidic Device
1Biomedical Engineering Department, Cornell University, 2Neurosurgical Laboratory for Translational Stem Cell Research, Weill Cornell Brain Tumor Center, Weill Cornell Medical College of Cornell University, 3Cell Morphology Department, Instituto de Investigacion Principe Felipe, 4Department of Chemical and Biomolecular Engineering, Cornell University
We demonstrate that the over expression of epidermal growth factor receptors (EGFR) enhances the motility of neural stem cells(NSCs) using a novel agarose gel based microfluidic device. This technology can be readily adaptable to other mammalian cell systems where cell sources are scarce, such as human neural stem cells, and the turn around time is critical.
JoVE General
Chemotactic Response of Marine Micro-Organisms to Micro-Scale Nutrient Layers
Environmental Microfluidics Group, MIT - Massachusetts Institute of Technology
The fabrication of microfluidic channels and their implementation in experiments for studying the chemotactic foraging behaviour of marine microbes within a patchy nutrient seascape and the swimming behaviour of bacteria within shear flow are described.
JoVE Bioengineering
Density Gradient Multilayered Polymerization (DGMP): A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering
1Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, 2Biomedical Sciences Program, University of California, San Diego, 3Department of Bioengineering, University of California, San Diego
Here we describe a unique strategy for creating biocompatible, layered matrices with continuous interfaces between distinct layers for tissue engineering. Such a scaffold could provide an ideal customizable environment to modulate cell behavior by various biological, chemical or mechanical cues
JoVE Neuroscience
Organotypic Slice Cultures of Embryonic Ventral Midbrain: A System to Study Dopaminergic Neuronal Development in vitro
Institute of Reconstructive Neurobiology, University of Bonn
A method to generate organotypic slices from the E12.5 murine embryonic midbrain is described. The organotypic slice cultures can be used to observe the behavior of dopaminergic neurons or other ventral midbrain neurons.
JoVE Immunology and Infection
Facilitating Drug Discovery: An Automated High-content Inflammation Assay in Zebrafish
1Institute for Toxicology and Genetics, Karlsruhe Institute of Technology, Karlsruhe, Germany, 2Institute for Applied Informatics, Karlsruhe Institute of Technology, Karlsruhe, Germany
Here we describe a novel high-content chemically induced inflammation assay aiming at the identification of immune-modulatory bioactives. We have successfully combined automated microscopy with custom developed software scripts enabling automated quantification of the inflammatory response as well as further data processing, analysis, mining, and storage.
JoVE Neuroscience
Automated Quantification of Synaptic Fluorescence in C. elegans
Department of Biological Sciences, University of Toledo
The abundance of neurotransmitter receptors clustered at synapses strongly influences synaptic strength. This method quantifies fluorescently-labeled neurotransmitter receptors in three dimensions with single-synapse resolution in C. elegans, allowing hundreds of synapses to be rapidly characterized within a single sample without distortions introduced by z-plane projection.
JoVE Immunology and Infection
Murine Model of CD40-activation of B cells
In this video, we demonstrate the procedure of CD40-activation and expansion of murine B cells from splenocytes of C57BL/6 mice, which can be used as a model antigen-presenting cell (APC) to study induction of immunity.
JoVE General
Studies of Bacterial Chemotaxis Using Microfluidics - Interview
Environmental Microfluidics Group, MIT - Massachusetts Institute of Technology
JoVE Immunology and Infection
Non-invasive Imaging of Leukocyte Homing and Migration in vivo
1Department of Pathology and Immunology, Washington University in St. Louis, 2National Institute of Neurological Disorders and Stroke, NINDS, NIH - National Institute of Health
Here, we describe a non-invasive two-photon (2P) microscopy approach to study leukocyte homing in the mouse footpad. We discuss the technical aspects of our tissue imaging preparation and walk the reader through a typical experiment from initial set up to execution and data collection.
JoVE Neuroscience
C. elegans Tracking and Behavioral Measurement
1Donnelly Centre, University of Toronto, 2Department of Physics and Astronomy, Vrije Universiteit, 3Okinawa Institute of Science and Technology, 4Department of Physics, University of Toronto
We have developed a video-rate tracking microscope system that can record and quantify C. elegans behavior at high resolution and high speeds. We have also developed computational methods to reduce the dimensionality of the worm images to a fundamental set of measurements that completely describe the shape of the worm.
JoVE General
Measuring Intracellular Ca2+ Changes in Human Sperm using Four Techniques: Conventional Fluorometry, Stopped Flow Fluorometry, Flow Cytometry and Single Cell Imaging
1Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología-Universidad Nacional Autónoma de México, 2Math and Sciences Department, Edison State College
Intracellular Ca2+ dynamics are very important in sperm physiology and Ca2+-sensitive fluorescent dyes constitute a versatile tool to study them. Population experiments (fluorometry and stopped flow fluorometry) and single cell experiments (flow cytometry and single cell imaging) are used to track spatio-temporal [Ca2+] changes in human sperm cells.
JoVE Neuroscience
Migratory Behavior of Cells Generated in Ganglionic Eminence Cultures
1Dept. of Anatomy, Physiology and Genetics, Uniformed Services University, 2Neuroscience Program, Uniformed Services University
Time lapse imaging of 3D tissue culture allows studying migratory behavior of individual cells originating from ganglionic eminence in reaction to fractionated protein extract from cerebral cortex.
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 Simple Protocol for Extracting Hemocytes from Wild Caterpillars
Department of Biological Sciences, The George Washington University
Insect hemocytes carry out many important functions, both immune and non-immune, throughout all stages of insect development. Our present knowledge of hemocyte types and function comes from studies on insect genetic models. Here, we present a method for extracting, quantifying and visualizing hemocytes from wild caterpillars.
JoVE Bioengineering
Cell Co-culture Patterning Using Aqueous Two-phase Systems
1Department of Biomedical Engineering, University of Michigan, 2Department of Macromolecular Science and Engineering, University of Michigan
Aqueous two-phase systems were used to simultaneously pattern multiple populations of cells. This fast and easy method for cell patterning takes advantage of the phase separation of aqueous solutions of dextran and polyethylene glycol and the interfacial tension that exists between the two polymer solutions.
JoVE General
Preparation of Quality Inositol Pyrophosphates
Inositol pyrophosphates play an important role in human pathologies such cancer, diabetes and obesity; however, the exact mechanism of action is a matter of dispute. The lack of commercially available inositol pyrophosphates renders detailed studies problematic. Here we describe a simple protocol to produce and isolate milligrams of inositol pyrophosphates.
JoVE Bioengineering
Three-dimensional Cell Culture Model for Measuring the Effects of Interstitial Fluid Flow on Tumor Cell Invasion
School of Biomedical Engineering, Science and Health Systems, Drexel University
Interstitial fluid flow is elevated in solid tumors and can modulate tumor cell invasion. Here we describe a technique to apply interstitial fluid flow to cells embedded in a matrix and then measure its effects on cell invasion. This technique can be easily adapted to study other systems.
JoVE Immunology and Infection
Infection of Zebrafish Embryos with Intracellular Bacterial Pathogens
1Department of Molecular Cell Biology, Institute of Biology, Leiden University, 2Department of Medical Microbiology and Infection Control, VU University Medical Center, 3Australian Regenerative Medicine Institute, Monash University
Transparent zebrafish embryos have proved useful model hosts to visualize and functionally study interactions between innate immune cells and intracellular bacterial pathogens, such as Salmonella typhimurium and Mycobacterium marinum. Micro-injection of bacteria and multi-color fluorescence imaging are essential techniques involved in the application of zebrafish embryo infection models.
JoVE Clinical and Translational Medicine
Evaluation of Cancer Stem Cell Migration Using Compartmentalizing Microfluidic Devices and Live Cell Imaging
1Department of Biomedical Engineering, University of Wisconsin-Madison, 2Materials Science Program, University of Wisconsin-Madison, 3Department of Neurological Surgery, University of Wisconsin-Madison, 4Carbone Comprehensive Cancer Center and Center for Stem Cell and Regenerative Medicine, University of Wisconsin-Madison
A compartmentalizing microfluidic device for investigating cancer stem cell migration is described. This novel platform creates a viable cellular microenvironment and enables microscopic visualization of live cell locomotion. Highly motile cancer cells are isolated to study molecular mechanisms of aggressive infiltration, potentially leading to more effective future therapies.
JoVE General
An In vitro FluoroBlok Tumor Invasion Assay
BD Biosciences, Discovery Labware
This video demonstrates how to measure cell invasion through cell culture inserts using a fluorescence-based methodology.
JoVE General
Profiling of Methyltransferases and Other S-adenosyl-L-homocysteine-binding Proteins by Capture Compound Mass Spectrometry (CCMS)
1Department of Biochemistry / Analytics, caprotec bioanalytics GmbH, 2Institute of Organic Chemistry, RWTH Aachen University
Capture Compounds are trifunctional small molecules to reduce the complexity of the proteome by functional reversible small molecule-protein interaction followed by photo-crosslinking and purification. Here we use a Capture Compound with S-adenosyl-L-homocysteine-binding as selectivity function to isolate methyltransferases from an Escherichia coli whole cell lysate and identify them by MS.
JoVE Immunology and Infection
Non-invasive Imaging of Disseminated Candidiasis in Zebrafish Larvae
Department of Molecular and Biomedical Sciences, University of Maine
The rapid development, small size and transparency of zebrafish are tremendous advantages for the study of innate immune control of infection1-4. Here we demonstrate techniques for infecting zebrafish larvae using the fungal pathogen Candida albicans by microinjection, methodology recently used to implicate phagocyte NADPH oxidase activity in control of fungal dimorphism5.
JoVE Neuroscience
Vertical T-maze Choice Assay for Arthropod Response to Odorants
Entomology and Nematology Department, University of Florida
A vertical, T-maze olfactometer is described for assaying the behavioral response of arthropods. The olfactometer allows the experimenter to measure choices performed by test subjects when subjected to two potential odor fields. Both attraction to and repulsion from odorants can be measured with this device.
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