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JoVE Science Education

General Laboratory Techniques

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Basic Methods in Cellular and Molecular Biology

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Model Organisms I

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Biophysics: The study of Physical phenomena and Physical processes as applied to living things.
 JoVE Bioengineering

Electric Cell-substrate Impedance Sensing for the Quantification of Endothelial Proliferation, Barrier Function, and Motility

1Department of Pulmonary Diseases, Institute for Cardiovascular Research, VU University Medical Center, 2Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center


JoVE 51300

This protocol reviews Electric Cell-substrate Impedance Sensing, a method to record and analyze the impedance spectrum of adherent cells for the quantification of cell attachment, proliferation, motility, and cellular responses to pharmacological and toxic stimuli. Detection of endothelial permeability and assessment of cell-cell and cell-substrate contacts are emphasized.

 JoVE Biology

Introduction to Solid Supported Membrane Based Electrophysiology

1Department of Biophysical Chemistry, Max Planck Institute of Biophysics, 2Buchmann Institute for Molecular Life Sciences, Goethe University Frankfurt


JoVE 50230

Here we present an electrophysiological method based on solid supported membranes with focus on its applications for the characterization of electrogenic membrane transporters.

 JoVE Applied Physics

Proton Transfer and Protein Conformation Dynamics in Photosensitive Proteins by Time-resolved Step-scan Fourier-transform Infrared Spectroscopy

1Experimental Molecular Biophysics, Freie Universität Berlin


JoVE 51622

Key steps of protein function, in particular backbone conformational changes and proton transfer reactions, often take place in the microsecond to millisecond time scale. These dynamical processes can be studied by time-resolved step-scan Fourier-transform infrared spectroscopy, in particular for proteins whose function is triggered by light.

 JoVE Biology

Measurement of Cellular Chemotaxis with ECIS/Taxis

1Molecular and Cell Biology, University of Connecticut, 2University of Connecticut


JoVE 3840

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 Bioengineering

Nanomanipulation of Single RNA Molecules by Optical Tweezers

1Nanoscale Engineering Graduate Program, College of Nanoscale Science and Engineering, University at Albany, State University of New York, 2Nanoscale Science Undergraduate Program, College of Nanoscale Science and Engineering, University at Albany, State University of New York, 3Nanobioscience Constellation, College of Nanoscale Science and Engineering, University at Albany, State University of New York, 4The RNA Institute, University at Albany, State University of New York, 5Department of Biological Sciences, University at Albany, State University of New York


JoVE 51542

Optical tweezers have been used to study RNA folding by stretching individual molecules from their 5’ and 3’ ends. Here common procedures are described to synthesize RNA molecules for tweezing, calibration of the instrument, and methods to manipulate single molecules.

 JoVE Bioengineering

Magnetic Tweezers for the Measurement of Twist and Torque

1Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology


JoVE 51503

Magnetic tweezers, a powerful single-molecule manipulation technique, can be adapted for the direct measurements of the twist (using a configuration called freely-orbiting magnetic tweezers) and torque (using a configuration termed magnetic torque tweezers) in biological macromolecules. Guidelines for performing such measurements are given, including applications to the study of DNA and associated nucleo-protein filaments.

 JoVE Bioengineering

Stretching Short Sequences of DNA with Constant Force Axial Optical Tweezers

1LSA Biophysics, University of Michigan, 2LSA Biophysics, Department of Physics, University of Michigan


JoVE 3405

We illustrate the use of a constant force axial optical tweezers to explore the mechanical properties of short DNA molecules. By stretching DNA axially, we minimize steric hindrances and artifacts arising in conventional lateral manipulation, allowing us to study DNA molecules as short as ~100 nm.

 JoVE Biology

Easy Measurement of Diffusion Coefficients of EGFP-tagged Plasma Membrane Proteins Using k-Space Image Correlation Spectroscopy

1Institute of Molecular Biology and Genetics and Interdisciplinary Nanoscience Center, Aarhus University, 2Departments of Chemistry and Physics, McGill University


JoVE 51074

This paper provides a step by step guide to the fluctuation analysis technique k-Space Image Correlation Spectroscopy (kICS) for measuring diffusion coefficients of fluorescently labeled plasma membrane proteins in live mammalian cells.

 JoVE Biology

Measuring Cation Transport by Na,K- and H,K-ATPase in Xenopus Oocytes by Atomic Absorption Spectrophotometry: An Alternative to Radioisotope Assays

1Institute of Chemistry, Technical University of Berlin, 2The Vollum Institute, Oregon Health & Science University


JoVE 50201

We describe a method to quantify the activity of K+-countertransporting P-type ATPases by heterologous expression of the enzymes in Xenopus oocytes and measuring Rb+ or Li+ uptake into individual cells by atomic absorption spectrophotometry. The method is a sensitive and safe alternative to radioisotope flux experiments facilitating complex kinetic studies.

 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


JoVE 3252

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 Chemistry

Determination of the Gas-phase Acidities of Oligopeptides

1Department of Chemistry, University of the Pacific


JoVE 4348

The determination of the gas-phase acidities of cysteine-containing oligopeptides is described. The experiments are performed using a triple quadrupole mass spectrometer. The relative acidities of the peptides are measured using collision-induced dissociation experiments, and the quantitative acidities are determined using the extended Cooks kinetic method.

 JoVE Biology

Isolation, Culture, and Functional Characterization of Adult Mouse Cardiomyoctyes

1Cardiovascular Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, 2Division of Cardiology, Sapienza University


JoVE 50289

Here we describe the isolation of adult mouse cardiomyoctyes using a Langendorff perfusion system. The resulting cells are Ca2+-tolerant, electrically quiescent and can be cultured and transfected with adeno- or lentiviruses to manipulate gene expression. Their functionality can also be analyzed using the MMSYS system and patch clamp techniques.

 JoVE Biology

One-channel Cell-attached Patch-clamp Recording

1Graduate Program in Neuroscience, SMBS, University at Buffalo, SUNY, 2Department of Biochemistry, SMBS, University at Buffalo, SUNY, 3Molecular and Cellular Neuroscience Department, The Scripps Research Institute, 4Department of Biochemistry and Graduate Program in Neuroscience, SMBS, University at Buffalo, SUNY


JoVE 51629

Described here is a procedure for obtaining long stretches of current recording from one ion channel with the cell-attached patch-clamp technique. This method allows for observing, in real time, the pattern of open-close channel conformations that underlie the biological signal. These data inform about channel properties in undisturbed biological membranes.

 JoVE Bioengineering

Helical Organization of Blood Coagulation Factor VIII on Lipid Nanotubes

1Department of Neuroscience and Cell Biology, University of Texas Medical Branch, 2Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, 3Sealy Center for Structural Biology and Molecular Biophysics, University of Texas Medical Branch


JoVE 51254

We present a combination of Cryo-electron microscopy, lipid nanotechnology, and structure analysis applied to resolve the membrane-bound structure of two highly homologous FVIII forms: human and porcine. The methodology developed in our laboratory to helically organize the two functional recombinant FVIII forms on negatively charged lipid nanotubes (LNT) is described.

 JoVE Neuroscience

In vivo Neuronal Calcium Imaging in C. elegans

1Department of Physiology and Biophysics, Boston University School of Medicine, 2Boston University Photonics Center


JoVE 50357

With its small transparent body, well-documented neuroanatomy and a host of amenable genetic techniques and reagents, C. elegans makes an ideal model organism for in vivo neuronal imaging using relatively simple, low-cost techniques. Here we describe single neuron imaging within intact adult animals using genetically encoded fluorescent calcium indicators.

 JoVE Bioengineering

Quantification of Global Diastolic Function by Kinematic Modeling-based Analysis of Transmitral Flow via the Parametrized Diastolic Filling Formalism

1Department of Biomedical Engineering, Washington University in St. Louis, 2Department of Physics, Washington University in St. Louis, 3Division of Biology and Biomedical Sciences, Washington University in St. Louis, 4Department of Medicine, Cardiovascular Division, Washington University in St. Louis, 5Cardiovascular Biophysics Lab, Washington University in St. Louis


JoVE 51471

Accurate, causality-based quantification of global diastolic function has been achieved by kinematic modeling-based analysis of transmitral flow via the Parametrized Diastolic Filling (PDF) formalism. PDF generates unique stiffness, relaxation, and load parameters and elucidates 'new' physiology while providing sensitive and specific indexes of dysfunction.

 JoVE Bioengineering

Measurement of Tension Release During Laser Induced Axon Lesion to Evaluate Axonal Adhesion to the Substrate at Piconewton and Millisecond Resolution

1Institute of Biophysics, National Research Council of Italy, 2Dipartimento di Sistemi e Informatica, Università di Firenze, 3Department of Neuroscience and Brain Technologies, Istituto Italiano di Tecnologia


JoVE 50477

We measured the tension release in an axon that was partially lesioned with a laser dissector by simultaneous force spectroscopy measurement performed on an optically-trapped probe adhered to the membrane of the axon. The developed experimental protocol evaluates the axon adhesion to the culture substrate.

 JoVE Bioengineering

Measuring the Mechanical Properties of Living Cells Using Atomic Force Microscopy

1Department of Physics, Worcester Polytechnic Institute, 2Department of Chemical Engineering, Worcester Polytechnic Institute


JoVE 50497

This paper demonstrates a protocol to characterize the mechanical properties of living cells by means of microindentation using an Atomic Force Microscope (AFM).

 JoVE Neuroscience

Manufacturing and Using Piggy-back Multibarrel Electrodes for In vivo Pharmacological Manipulations of Neural Responses

1Department of Physiology & Biophysics, University of Colorado Medical Campus


JoVE 4358

Iontophoresis of neural agonists and antagonists during extracellular in vivo recordings is a powerful way to manipulate a neuron’s microenvironment. These manipulations can most easily be done via piggy-back multibarrel electrodes. Here we describe how to manufacture them and use them during auditory recordings.

 JoVE Bioengineering

Flexural Rigidity Measurements of Biopolymers Using Gliding Assays

1Department of Physics, Lawrence University


JoVE 50117

A method to measure the persistence length or flexural rigidity of biopolymers is described. The method uses a kinesin-driven microtubule gliding assay to experimentally determine the persistence length of individual microtubules and is adaptable to actin-based gliding assays.

 JoVE Bioengineering

Correlative Microscopy for 3D Structural Analysis of Dynamic Interactions

1Department of Structural Biology, University of Pittsburgh School of Medicine, 2Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine


JoVE 50386

We describe a correlative microscopy method that combines high-speed 3D live-cell fluorescent light microscopy and high-resolution cryo-electron tomography. We demonstrate the capability of the correlative method by imaging dynamic, small HIV-1 particles interacting with host HeLa cells.

 JoVE Biology

Single-molecule Imaging of Gene Regulation In vivo Using Cotranslational Activation by Cleavage (CoTrAC)

1Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, 2Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, 3Department of Physics, Jilin University


JoVE 50042

We describe a fluorescence microscopy method, Co-Translational Activation by Cleavage (CoTrAC), to image the production of protein molecules in live cells with single-molecule precision without perturbing the protein's functionality. This method has been used to follow the stochastic expression dynamics of a transcription factor, the λ repressor CI 1.

 JoVE Immunology and Infection

Isolation of Adipose Tissue Immune Cells

1Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine


JoVE 50707

Adipose tissue (AT) is a site of intense immune cell activation and interaction. Almost all cells of the immune system are present in AT and their ratios are altered by obesity. Proper isolation, quantification, and characterization of AT immune cell populations are critical for understanding their role in immunometabolic disease.

 JoVE Biology

Giant Liposome Preparation for Imaging and Patch-Clamp Electrophysiology

1Department of Physiology and Biophysics, University of Washington


JoVE 50227

Reconstituting functional membrane proteins into giant liposomes of defined composition is a powerful approach when combined with patch-clamp electrophysiology. However, conventional giant liposome production may be incompatible with protein stability. We describe protocols for producing giant liposomes from pure lipids or small liposomes containing ion channels.

 JoVE Biology

Super-resolution Imaging of the Bacterial Division Machinery

1Department of Biophysics and Biophysical Chemistry, The Johns Hopkins University School of Medicine


JoVE 50048

We describe a super-resolution imaging method to probe the structural organization of the bacterial FtsZ-ring, an essential apparatus for cell division. This method is based on quantitative analyses of photoactivated localization microscopy (PALM) images and can be applied to other bacterial cytoskeletal proteins.

 JoVE Biology

Fluorescence-based Measurement of Store-operated Calcium Entry in Live Cells: from Cultured Cancer Cell to Skeletal Muscle Fiber

1Department of Physiology and Biophysics, Confocal Microscopy and Cell Imaging Core, Robert Wood Johnson Medical School, 2Department of Physiology and Biophysics, Robert Wood Johnson Medical School, 3Muscle Biology Research Group-MUBIG Schools of Nursing & Medicine, University of Missouri-Kansas City


JoVE 3415

The extent of store-operated Ca2+ entry (SOCE) can be monitored using fluorescent Ca2+ indicators. Mn2+ quenching of such indicators assays SOCE in cultured cells and skeletal muscle fibers. A technique allowing spatial and temporal resolution of SOCE by confocal imaging of mechanically skinned muscle fibers is also described.

 JoVE Applied Physics

Construction of a High Resolution Microscope with Conventional and Holographic Optical Trapping Capabilities

1Department of Physics and Astronomy, University of Utah


JoVE 50481

The system described herein employs a traditional optical trap as well as an independent holographic optical trapping line, capable of creating and manipulating multiple traps. This allows for the creation of complex geometric arrangements of refractive particles while also permitting simultaneous high-speed, high-resolution measurements of the activity of biological enzymes.

 JoVE Biology

Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules

1Department of Chemical and Biological Engineering, Princeton University


JoVE 50476

We developed computational de novo protein design methods capable of tackling several important areas of protein design. To disseminate these methods we present Protein WISDOM, an online tool for protein design (http://www.proteinwisdom.org). Starting from a structural template, design of monomeric proteins for increased stability and complexes for increased binding affinity can be performed.

 JoVE Bioengineering

Visualizing Proteins and Macromolecular Complexes by Negative Stain EM: from Grid Preparation to Image Acquisition

1Graduate Group in Biophysics, University of California San Francisco, 2Department of Biochemistry and Biophysics, University of California San Francisco


JoVE 3227

Visualizing protein samples by negative stain electron microscopy (EM) has become a popular structural analysis method. It is useful for quantitative structural analysis, such as calculating a 3D reconstruction of the molecules being studied, and also for qualitative examination of the quality of protein preparations. In this article we present detailed protocols for preparing the EM grids, staining the sample and visualizing the sample in an electron microscope. Novice users can follow these protocols easily and to utilize negative stain EM as a routine assay, in addition to other biochemical assays, for evaluating their protein samples.

 JoVE Biology

Isolation and Biophysical Study of Fruit Cuticles

1Department of Chemistry, City College of New York, City University of New York Graduate Center and Institute for Macromolecular Assemblies, 2Department of Chemical Engineering, City College of New York


JoVE 3529

Aerial plant organs are protected by the cuticle, a supramolecular biopolyester-wax assembly. We present protocols to monitor selective removal of epi- and intracuticular waxes from tomato fruit cuticles on molecular and micro scales by solid-state NMR and atomic force microscopy, respectively, and to assess the cross-linking capacity of engineered cuticular biopolyesters.

 JoVE Clinical and Translational Medicine

MRI-guided Disruption of the Blood-brain Barrier using Transcranial Focused Ultrasound in a Rat Model

1Imaging Research, Sunnybrook Research Institute, 2Department of Medical Biophysics, University of Toronto, 3Department of Medical Biophysics, and Institute of Biomaterials & Biomedical Engineering (IBBME), University of Toronto


JoVE 3555

Microbubble-mediated focused ultrasound disruption of the blood-brain barrier (BBB) is a promising technique for non-invasive targeted drug delivery in the brain1-3. This protocol outlines the experimental procedure for MRI-guided transcranial BBB disruption in a rat model.

 JoVE Biology

Determination of Protein-ligand Interactions Using Differential Scanning Fluorimetry

1Department of Biosciences, University of Exeter


JoVE 51809

Differential scanning fluorimetry is a widely used method for screening libraries of small molecules for interactions with proteins. Here, we present a straightforward method to extend these analyses to provide an estimate of the dissociation constant between a small molecule and its protein partner.

 JoVE Bioengineering

Small and Wide Angle X-Ray Scattering Studies of Biological Macromolecules in Solution

1Department of Mechanical, Aerospace, and Nuclear Engineering, Rensselaer Polytechnic Institute


JoVE 4160

The demonstration of the small and wide angle X-ray scattering (SWAXS) procedure has become instrumental in the study of biological macromolecules. Through the use of the instrumentation and procedures of specific angle methods and preparation, the experimental data from the SWAXS displays the atomic and nano-scale characterization of macromolecules.

 JoVE Clinical and Translational Medicine

Molecular Imaging to Target Transplanted Muscle Progenitor Cells

1Imaging Program, Lawson Health Research Institute, 2Department of Anatomy and Cell Biology, Western University, 3Department of Medical Biophysics, Western University


JoVE 50119

A non-invasive means to evaluate the success of myoblast transplantation is described. The method takes advantage of a unified fusion reporter gene composed of genes whose expression can be imaged with different imaging modalities. Here, we make use of a fluc reporter gene sequence to target cells via bioluminescence imaging.

 JoVE Biology

Modeling Biological Membranes with Circuit Boards and Measuring Electrical Signals in Axons: Student Laboratory Exercises

1Department of Biology, University of Kentucky, 2Department of Physiology, University of Toronto


JoVE 2325

This is a demonstration of how biological membranes can be understood using electrical models. We also demonstrate procedures for recording action potentials from the ventral nerve cord of the crayfish for student orientated laboratories.

 JoVE Biology

Test Samples for Optimizing STORM Super-Resolution Microscopy

1Analytical Science Division, National Physical Laboratory


JoVE 50579

We describe the preparation of three test samples and how they can be used to optimize and assess the performance of STORM microscopes. Using these examples we show how to acquire raw data and then process it to acquire super-resolution images in cells of approximately 30-50 nm resolution.

 JoVE Biology

Reconstitution of a Kv Channel into Lipid Membranes for Structural and Functional Studies

1Department of Cell Biology, University of Texas Southwestern Medical Center at Dallas


JoVE 50436

Procedures for complete reconstitution of a prototype voltage-gated potassium channel into lipid membranes are described. The reconstituted channels are suitable for biochemical assays, electrical recordings, ligand screening and electron crystallographic studies. These methods may have general applications to the structural and functional studies of other membrane proteins.

 JoVE Biology

The Xenopus Oocyte Cut-open Vaseline Gap Voltage-clamp Technique With Fluorometry

1Department of Biomedical Engineering, Washington University in St. Louis


JoVE 51040

The cut-open Vaseline gap approach is used to obtain low noise recordings of ionic and gating currents from voltage-dependent ion channels expressed in Xenopus oocytes with high resolution of fast channel kinetics. With minor modification, voltage clamp fluorometry can be coupled to the cut-open oocyte protocol.

 JoVE Bioengineering

Self-reporting Scaffolds for 3-Dimensional Cell Culture

1School of Molecular Medical Sciences, University of Nottingham, 2Division of Drug Delivery and Tissue Engineering, University of Nottingham, 3Laboratory of Biophysics and Surface Analysis, University of Nottingham


JoVE 50608

Biocompatible pH responsive sol-gel nanosensors can be incorporated into poly(lactic-co-glycolic acid) (PLGA) electrospun scaffolds. The produced self-reporting scaffolds can be used for in situ monitoring of microenvironmental conditions whilst culturing cells upon the scaffold. This is beneficial as the 3D cellular construct can be monitored in real-time without disturbing the experiment.

 JoVE Neuroscience

Deriving the Time Course of Glutamate Clearance with a Deconvolution Analysis of Astrocytic Transporter Currents

1Synaptic Physiology Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health


JoVE 50708

We describe an analytical method to estimate the lifetime of glutamate at astrocytic membranes from electrophysiological recordings of glutamate transporter currents in astrocytes.

 JoVE Biology

Live Cell Imaging of Primary Rat Neonatal Cardiomyocytes Following Adenoviral and Lentiviral Transduction Using Confocal Spinning Disk Microscopy

1Max-Planck-Institute for Molecular Biomedicine and Institute of Cell Biology, 2Department of Internal Medicine, Yale Cardiovascular Research Center and Section of Cardiovascular Medicine


JoVE 51666

This protocol describes a method of live cell imaging using primary rat neonatal cardiomyocytes following lentiviral and adenoviral transduction using confocal spinning disk microscopy. This enables detailed observations of cellular processes in living cardiomyocytes.

 JoVE Clinical and Translational Medicine

Technique and Considerations in the Use of 4x1 Ring High-definition Transcranial Direct Current Stimulation (HD-tDCS)

1Laboratory of Neuromodulation, Department of Physical Medicine & Rehabilitation, Spaulding Rehabilitation Hospital and Massachusetts General Hospital, Harvard Medical School, 2School of Medicine, Pontifical Catholic University of Ecuador, 3Charité University Medicine Berlin, 4The City College of The City University of New York, 5Headache & Orofacial Pain Effort (H.O.P.E.), Biologic & Materials Sciences, School of Dentistry, University of Michigan


JoVE 50309

High-definition transcranial direct current stimulation (HD-tDCS), with its 4x1-ring montage, is a noninvasive brain stimulation technique that combines both the neuromodulatory effects of conventional tDCS with increased focality. This article provides a systematic demonstration of the use of 4x1 HD-tDCS, and the considerations needed for safe and effective stimulation.

 JoVE Bioengineering

Luminescence Resonance Energy Transfer to Study Conformational Changes in Membrane Proteins Expressed in Mammalian Cells

1Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston


JoVE 51895

We describe here an improved Luminescence Resonance Energy Transfer (LRET) method where we introduce a protease cleavage site between the donor and acceptor fluorophore sites. This modification allows us to obtain specific LRET signals arising from membrane proteins of interest, allowing for the study of membrane proteins without protein purification.

 JoVE Biology

Fluorescence Imaging with One-nanometer Accuracy (FIONA)

1Department of Physics, University of Illinois at Urbana-Champaign, 2Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, 3Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign


JoVE 51774

Single fluorophores can be localized with nanometer precision using FIONA. Here a summary of the FIONA technique is reported, and how to carry out FIONA experiments is described.

 JoVE Biology

Acquiring Fluorescence Time-lapse Movies of Budding Yeast and Analyzing Single-cell Dynamics using GRAFTS

1Department of Chemical Engineering, Massachusetts Institute of Technology


JoVE 50456

We present a simple protocol to obtain fluorescence microscopy movies of growing yeast cells, and a GUI-based software package to extract single-cell time series data. The analysis includes automated lineage and division time assignment integrated with visual inspection and manual curation of tracked data.

 JoVE Biology

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


JoVE 50344

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 Bioengineering

A Novel Three-dimensional Flow Chamber Device to Study Chemokine-directed Extravasation of Cells Circulating under Physiological Flow Conditions

1Torrey Pines Institute for Molecular Studies, 2Cascade LifeSciences Inc.


JoVE 50959

The three-dimensional flow chamber device is a novel in vitro technology for the quantitative and step-wise evaluation of the extravasation cascade of cells circulating under conditions of physiological shear stress. The device therefore fills a critical need for basic, preclinical, and clinical studies of cell migration.

 JoVE Neuroscience

Fast Micro-iontophoresis of Glutamate and GABA: A Useful Tool to Investigate Synaptic Integration

1NRW Research Group 'Dendritic integration in the CNS', Department of Epileptology, University of Bonn, 2Neuronal Networks Group, Deutsches Zentrum für Neurodegenerative Erkrankungen e.V. (DZNE)


JoVE 50701

In this article we introduce fast micro-iontophoresis of neurotransmitters as a technique to investigate integration of postsynaptic signals with high spatial and temporal precision.

 JoVE Neuroscience

Preparation of Primary Neurons for Visualizing Neurites in a Frozen-hydrated State Using Cryo-Electron Tomography

1Department of Molecular Physiology and Biophysics, Baylor College of Medicine, 2Department of Neuroscience, Baylor College of Medicine, 3Department of Neuroscience, University of California at San Diego, 4National Center for Macromolecular Imaging, Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine


JoVE 50783

To preserve neuronal processes for ultrastructural analysis, we describe a protocol for plating of primary neurons on electron microscopy grids followed by flash freezing, yielding samples suspended in a layer of vitreous ice. These samples can be examined with a cryo-electron microscope to visualize structures at the nanometer scale.

 JoVE Environment

Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy

1Lawrence Berkeley National Laboratory, The Molecular Foundry


JoVE 51087

More than half of proteins are small proteins (molecular mass <200 kDa) that are challenging for both electron microscope imaging and three-dimensional reconstructions. Optimized negative staining is a robust and high-throughput protocol to obtain high contrast and relatively high resolution (~1 nm) images of small asymmetric proteins or complexes under different physiological conditions.

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