<h2 class="jove_title"><a href="https://www.jove.com/video/3505/">Measuring Fast Calcium Fluxes in Cardiomyocytes</a></h2>
Urszula Golebiewska1, Suzanne Scarlata2 1Department of Biological Sciences and Geology, Queensborough Community College, 2Department of Physiology &#38; Biophysics, Stony Brook University
We present a method to isolate rapid (microsecond) calcium events from slower fluxes in living cells using laser scanning confocal microscopy. The method measures fluorescence intensity fluctuations of calcium indicators by recording line scans of several hundred pixels in a cell. Histogram analysis allows us to isolate the time scales of different calcium fluxes.
<h2 class="jove_title"><a href="https://www.jove.com/video/3373/">Solid-phase Submonomer Synthesis of Peptoid Polymers and their Self-Assembly into Highly-Ordered Nanosheets	</a></h2>
Helen Tran, Sarah L. Gael, Michael D. Connolly, Ronald N. Zuckermann Molecular Foundry, Lawrence Berkeley National Laboratory
A simple and general manual peptoid synthesis method involving basic equipment and commercially available reagents is outlined, enabling peptoids to be easily synthesized in most laboratories. The synthesis, purification and characterization of an amphiphilic peptoid 36mer is described, as well as its self-assembly into highly-ordered nanosheets.
<h2 class="jove_title"><a href="https://www.jove.com/video/3258/">Behavioral Assessment of Manual Dexterity in Non-Human Primates</a></h2>
Eric Schmidlin*, M&#233;lanie Kaeser*, Anne- Dominique Gindrat, Julie Savidan, Pauline Chatagny, Simon Badoud, Adjia Hamadjida, Marie-Laure Beaud, Thierry Wannier, Abderraouf Belhaj-Saif, Eric M. Rouiller Department of Medicine, University of Fribourg * These authors contributed equally
As manual dexterity is a prerogative mainly of primates, behavioral tasks have been developed in macaque monkeys. Four reach and grasp prehension tasks, measuring hand manipulation ability and force, allow to establish functional recovery after a lesion of the central nervous system and to test the effect of a treatment.
<h2 class="jove_title"><a href="https://www.jove.com/video/3588/">Determining the Phagocytic Activity of Clinical Antibody Samples</a></h2>
Elizabeth G. McAndrew1, Anne-Sophie Dugast1, Anna F. Licht1, Justin R. Eusebio1, Galit Alter1, Margaret E. Ackerman2 1Massachusetts General Hospital, Ragon Institute of MGH, MIT, and Harvard, 2Thayer School of Engineering, Dartmouth College
We present a high-throughput flow cytometric assay to determine the phagocytic activity of antigen-specific antibodies from clinical samples, utilizing fluorescent antigen-coated beads and a monocytic cell line expressing multiple Fc receptors-providing receptor usage and phagocytic activity determinations in a standardized and reproducible fashion for any antigen of interest.
<h2 class="jove_title"><a href="https://www.jove.com/video/3187/">Study of the Actin Cytoskeleton in Live Endothelial Cells Expressing GFP-Actin</a></h2>
Travis M. Doggett, Jerome W. Breslin Department of Physiology, Louisiana State University Health Sciences Center	Microscopic imaging of live endothelial cells expressing GFP-actin allows characterization of dynamic changes in cytoskeletal structures. Unlike techniques that use fixed specimens, this method provides a detailed assessment of temporal changes in the actin cytoskeleton in the same cells before, during, and after various physical, pharmacological, or inflammatory stimuli.
<h2 class="jove_title"><a href="https://www.jove.com/video/3260/">Implantation of Radiotelemetry Transmitters Yielding Data on ECG, Heart Rate, Core Body Temperature and Activity in Free-Moving Laboratory Mice</a></h2>
Nikola Cesarovic1, Paulin Jirkof2, Andreas Rettich2, Margarete Arras1 1Division of Surgical Research, University Hospital Zurich, 2Institute of Laboratory Animal Science, University of Zurich
A surgical technique for implantation of commercially available telemetry transmitters used for continuous measurement of biopotential (one-lead ECG), heart rate, core body temperature and locomotor activity in freely moving mice is shown. Recommendations and protocols for post-operative care and pain relief, improving recovery, wellbeing and survival rate are also presented.
<h2 class="jove_title"><a href="https://www.jove.com/video/3381">Electroporation of Craniofacial Mesenchyme</a></h2>
Jacqueline M. Tabler, Karen J. Liu Department of Craniofacial Development, King&#39;s College London
Craniofacial cartilages develop in close contact with other tissues and are difficult to manipulate in live animals. We are using electroporation to deliver molecular tools during growth of the craniofacial skeleton while bypassing early embryonic effects. This approach will allow us to efficiently test candidate molecules in vivo.
<h2 class="jove_title"><a href="https://www.jove.com/video/3379">Slice Electrophysiological Recording and Optogenetic Stimulation of Adult-Born Neurons in the Olfactory Bulb</a></h2>
Matthew Valley, Sebastian Wagner, Benjamin W. Gallarda, Pierre-Marie Lledo Laboratory for Perception and Memory, Institut Pasteur and Centre National de la Recherche Scientifique (CNRS)
Adult-born neurons expressing ChR2 can be manipulated in slice electrophysiological preparations in order to examine their contribution towards the function of olfactory neural circuits.

november 2011: this month in jove

The November issue of the Journal of Visualized Experiments begins with a series of behavioral experiments for measuring prehension ability in primates. Investigators from the University of Fribourg demonstrate four grasping and reaching tasks, which all follow common steps for comfortably restraining the animal and correctly positioning its head. The Brinkman Board test is used to assess reaching for static objects, while the Rotating Brinkman Board presents a condition where animals need to anticipate object position. The Brinkman Box test allows dexterity to be investigated under conditions where the monkey has fewer degrees of freedom to perform grasping task and cannot use vision to guide movement. While the brinkman board and box tests solely rely on analysis of video obtained during experiments, in order to generate data, the drawer reaching task apparatus has detectors that monitor grip and load forces as well as drawer position, making this test appropriate for measuring strength as well as prehension ability. In combination, these four tests provide a powerful approach to assess motor control in the distal forelimb, and the Fribourg group presents data from both lesion studies and experiments aimed to test potential therapies for spinal cord injury. In Clinical and Translational Medicine, collaborators from the Division of Surgical Research at the University Hospital Zurich and the Institute of Laboratory Animal Science at the University of Zurich, work together to implant a radiotelemetry device in mice, which can be used to take physiological measurements in freely moving animals. The devices consist of two leads, which are capable of measuring biopotential activity (ECG, EEG, or EMG) as well as temperature, and a lightweight radio transmitter (1-200 Hz in bandwidth) that can be implanted in the abdominal cavity. The stress of animal handling as well as anesthesia are known confounding elements, which impact physiological measurements, so radiotelemetry devices are a convenient way to acquire physiological data from animals in an unperturbed, natural state, as well as take measurements of multiple animals, simultaneously. JoVE also adds to its host of electroporation protocols with a method for introducing macromoledules into the craniofacial mesenchyme of developing Xenopus laevis tadpoles, from the Liu Lab at King's College London. Xenopus tadpoles are transferred to an electroporation chamber, where they can be injected with nucleic acid, and then subjected to electroporation, which will deliver expression vectors or morpholinos into the developing craniofacial tissue. Gene delivery into differentiated cartilage is inherently problematic, and the Liu lab shows that, through elecroporation, morpholinos can be introduced into procartilaginous and cartilaginous tissues of the Xenopus tadpole throughout a range of developmental stages. In Bioengineering, the Zuckerman group from the Lawrence Berkeley National Laboratory illustrates their method for synthesizing peptoids, a class of biologically-active organic molecules that resemble peptides, but have their side chains attached to the nitrogen of the peptide bond, rather than the alpha carbon, which makes them resistant to proteolysis. The Zuckerman group demonstrates submonomer synthesis, where peptide monomers are added sequentially in bromoacetylation and bromide displacement reactions, allowing peptoid chain length and composition to be precisely controlled. Furthermore, the fact that synthesis is done on amide resin, and is hence, solid phase synthesis, greatly simplifies purification of the synthesized peptoids. These authors also show how the newly-synthesized peptoids can be induced to self-assemble into highly-ordered structures, termed nanosheets, which can be visualized by fluorescence and scanning electron microscopy. These nanosheets could serve as a potential platform for protein and membrane mimetics. Later in the month, JoVE features an assay for measuring phagocytic activity in cells exposed to clinical antibody samples for our Immunology and Infection section. Collaborators from Massachusetts General Hospital and Dartmouth College show that THP-1 cells will engulf antibody-bound antigen coated beads in timelapse video microscopy and present a plate-based flow cytometry method for quantifying the extent of phagocytosis. Our authors use this method to determine the ability of different antibody samples to induce phagocytosis, such as those from HIV patients both treated and untreated with anti-retroviral therapies. This summary of JoVE's content in November provides a brief overview of JoVE's highlights for the month. Other noteworthy articles include methods for live imaging of cytoskeletal dynamics in endothelial cells, measuring calcium flux in cardiomyocytes, and slice electrophysiology of adult born neurons in the olfactory bulb.

Watch this Scientific Journal Video about November 2011: This Month in JoVE at JoVE.com

November 2011: This Month in JoVE

Dieses November in JoVE

The November issue of the Journal of Visualized Experiments begins with a series of behavioral experiments for measuring prehension ability in primates. ...

Research

JoVE Journal

Biology

Serie: November 2011: This Month in JoVE

December 2011: This Month in JoVE

The Journal of Visualized Experiments (JoVE) closes 2011 with a December issue that begins by honoring World's AIDS Day with an article that deals with visualizing the structure of HIV envelope glycoproteins, which are a critical component of the viral infection pathway.
One key method for determining virion envelope structure is cryo-electron tomography. Prior to using this technique, soluble virus must be vitrified on a gold grid - a process that involves adding gold to a viral suspension, transferring that suspension to the grid, and plunging the grid into liquid ethane. The virus particles, present in a thin film of liquid atop the grid, are frozen at a rate greater than 100,000 K per second. Following vitrification, collected samples are loaded into the electron microscope and precisely defined regions on the grid are hit with a focused electron beam, while the grid is tilted. The tilt series that results from these imaging sessions is comprised of hundreds of individual images, which must be aligned and averaged to resolve individual envelope proteins. This computationally intense process often requires computer clusters, capable of remote-access parallel processing, like NIH's Biowulf.
Once completed, 3D models can not only provide insight into the structure of envelope glycoproteins themselves, but can also reveal the manner in which they interact other proteins, like the neutralizing antibody B12, which is information that can greatly influence vaccine design.
This article, from the laboratory of Dr. Sriram Subramaniam involves contributions from twenty authors, including not only graduates students and postdocs, but also middle and highschool students, showing that you are never too young to make an impact in the fight against a disease that has infected over 60 million people, worldwide.
In Clinical and Translational Medicine, JoVE presents an article from the University College London and the University of Oxford that investigates the physiological correlates of nociception, or pain sensation, in infants. Typically, infants respond to noxious stimuli by crying and changing their facial expressions - reactions considered to be autonomic and reliant on subcortical regions of the brain.
In order to understand the involvement of the cerebral cortex and spinal cord in infant nociception our authors first prepare infants for EEG and EMG recordings. They also set up physiological monitoring and video recording, so that the infant's facial expressions can be linked to physiological data. Following assessment of infant responses to neutral tactile stimuli, the baby is subjected to heel lancing - a clinical procedure used to obtain blood samples from infants. After data processing, electrical potentials can be compared between noxious and non-noxious stimuli in these infants, which will hopefully lead to meaningful conclusions, regarding how nociceptive information is interpreted by newborns.
JoVE's Bioengineering section takes a more clinical focus in December with an article from the University of Wisconsin, Madison, which involves the fabrication of a compartmentalized microfluidic device for studying cancer stem cell migration. Following device design, master molds are created by photolithography for the application of poly dimethyl siloxane (PDMS)- an organosilicon compound that is transparent and biologically inert - making it suitable for microfluidics applications with live cells. Specifically, our authors load tumor stem cells derived from multiform glioblastoma, a highly malignant brain cancer, into the compartmentalized devices and then place them in a specialized imaging system that acquires data over a few days. 
Time lapse video from these authors show that cancer stem cells are able to regulate their morphology in the size-constrained microfluidic channels much like they would in the interstitial space, and fluidic isolation made possible by chambers like these facilitates drug screening for potential therapeutics that can affect cancer stem cell migration.
Late on in the month, JoVE travels to France, where investigators from the Institute Pasteur and the Centre National de la Recherche Scientifique, illustrate a method for inducing expression of channel-rhodopsin, a light-activated ion channel, into the olfactory bulb of mice, as well as subsequent neuronal stimulation using a miniature light emitting diode (LED).
Our authors give detailed step-by-step instructions for preparing injection needles, and for precise injection of virus into the rostral migratory stream, a site of migration for adult born neuroblasts. Furthermore, they show how miniature LED stimulation devices are fabricated, calibrated, and implanted atop a cranial window prepared over the bulb. In particular, our authors are interested in the way that adult born neurons integrate into existing olfactory neuronal circuits, and they show that LED stimulation in awake behaving animals, can lead to activation of virally-tranduced neuroblasts in the olfactory bulb, which have migrated from the rostral migratory stream.
These video-articles comprise JoVE's highlights for the month of December. Other noteworthy upcoming articles illustrate methods for patterning conductive inks into microelectrodes, measuring cytosolic calcium in contractile lymphatics, and imaging neuronal responses to pheromones in the vomeronasal organ.

Here are some highlights from the December 2011 Issue of Journal of Visualized Experiments (JoVE).

Im Dezember dieses Jahres in JoVE

The Journal of Visualized Experiments (JoVE) closes 2011 with a December issue that begins by honoring World's AIDS Day with an article that deals with ...

Forschung

Biologie

August 2012: This Month in JoVE

Traditional microscopy requires lens objectives to magnify specimens, and can involve numerous optical components like additional objectives, filters, and mirrors to refract and direct light to optical sensors. The August 2012 issue of JoVE (Journal of Visualized Experiments) is marked by the third publication from the Ozcan Lab (University of California, Los Angeles) on their lens free lens-free "on-chip" microscopy platform, which they have pioneered.
Back in 2009, the Ozcan lab published their first article, which demonstrated the use of a lens-free imaging platform that could image cells within whole blood. This imaging technique involves placing a sample directly on an optoelectric sensor array, like the CCD or CMOS chips found in the standard camcorder. Then, LED illumination from above the sample casts shadows on the chip, which are recorded as digital holograms that can be subjected to image processing. Not only does this lensless imaging system allow for an extremely large field of view (FOV of 0.6-8 cm2), the lack of bulky optical components makes it excellent for microfluidic applications. The Ozcan group demonstrates this in their follow-up 2011 JoVE article, in which fluorescently labeled white blood cells are imaged within a microfluidic chip.
This August in JoVE, the Ozcan group demonstrates further advancements to their lens-free microscopy platform by applying it to tomographic imaging of the nematode, C. elegans. By changing the angle of illumination, using a motorized rotation stage, the Ozcan group shows how multiple holographic slices may be generated for a sample-which, following processing, can be combined into tomograms. The Ozcan group confirms that their 3D reconstruction of the worm is successful by citing that the pharyngeal tube, a 5-micron-thick structure, is only visible in a single slice and in a 3D reconstructed cross section.
The compact and inexpensive nature of the lens-free imaging systems - designed and assembled by the Ozcan group - make them well suited for point-of-care diagnostic applications, or even cell phone based microscopy that can be performed in the field.
At the Ernest Gallo Clinic and Research Center at University of California, San Francisco JoVE learns how to visualize morphological changes in single cells using three-dimensional fluorescence microscopy and computer analysis. In this procedure, cells are given agonist, antagonist plus agonist, or no treatment, and prepared for immunofluorescence microscopy. Three-dimensional images are acquired with the help of a two-photon laser system, and the fluorescence data are processed on the computer using Imaris and Matlab.
To demonstrate the power of this technique, our authors quantify single-cell phenotypic changes in transfected HEK293 cells. Three-dimensional morphometric analysis shows dramatic changes in cells treated with agonist. These changes are not always apparent when using conventional 2D analysis. This technique allows morphological changes in single cells to be quantified and characterized in response to pharmacologic stimuli, offering a powerful new assessment tool to the arsenal of drug discovery research.
This month, in JoVE Applied Physics, we travel to South Carolina to visit the School of Materials Science and Engineering at Clemson University. Here, researchers show us how to construct lithium batteries for laboratory research.
The basic components of a lithium coin cell are the working electrode, the lithium electrolyte, and a lithium foil counter electrode. The different components of the coin cell are carefully assembled and sealed with a crimping machine. The finished coin cell can then be characterized in a battery tester. Using their homemade coin cells, our researchers can experiment with using different materials to make the cathodes of lithium coin cells.
This month in JoVE Immunology and Infection, we head to Basel, Switzerland to witness two techniques for diagnosing helminth infections. We also see how investigators perform the shuttle run test in children, in order to assess the effects of these infections on physical fitness.
Helminths are parasitic worms that can live inside humans and animals. Many species infect the intestinal tract, and may be transmitted through soil. Our authors demonstrate the Kato-Katz technique, which is used for diagnosing several common soil-transmitted helminths. They also demonstrate the Baermann technique, which uses light to stimulate the migration of certain types of phototactic helminths out of sample preparations. The various types of helminth species can subsequently be determined based on morphology.
Our authors then demonstrate how to set up and perform a 20-meter shuttle run test for children, and to measure the impact of helminth infections on physical fitness. This method can help determine the true burden of soil-transmitted helminth infections on overall health.
This brief summary highlights but a few notable video-articles that will be released in August. We'll also feature articles that demonstrate how to express genes in the midbrain of the chick embryo, study neutrophil transmigration in parallel flow chambers lined with endothelial cells, and precisely measure proteolytic degradation of the extracellular matrix.

Traditional microscopy requires lens objectives to magnify specimens, and can involve numerous optical components like additional objectives, filters, and mirrors to refract and direct light to optical sensors. The August 2012 issue of JoVE (Journal of Visualized Experiments) is marked by the third publication from the Ozcan Lab (University of California, Los Angeles) on their lens-free "on-chip" microscopy platform, which they have pioneered.

August 2012: Diesen Monat in JoVE

Traditional microscopy requires lens objectives to magnify specimens, and can involve numerous optical components like additional objectives, filters, and ...

2012: A Year In Review

Here's a look at some of the milestones and highlights of the year 2012 in Journal of Visualized Experiments (JoVE).
At the start of 2012, JoVE reached a major milestone by publishing our 1500th article. In this publication, in JoVE Bioengineering, Yu et al. described a method for analyzing the free radical composition of cigarette smoke. This method simulated cigarette puffing with a single-port smoking device, and combined it with electron spin resonance (ESR) spectroscopy to measure the degree to which antioxidant compounds can scavenge free radicals in cigarette smoke. A possible outcome of this research is the creation of less harmful cigarettes.
JoVE Bioengineering also contained the most-viewed article of 2012. Owczarczak et al. described a method for hacking a standard inkjet printer, loading ink cartridges with a cell-suspension "bioink," and printing lines of live cells onto glass slides. The thermal inkjet printing process creates temporary pores in the cell membrane, visualized by the incorporation of fluorescently labeled actin monomers. This concept, called "bioprinting," has many potential applications in cell and tissue engineering.
Also in 2012, JoVE Bioengineering featured a protocol by Hsia et al. for isolating and purifying spider silk protein, spinning it into fibers, and assessing the fiber strength. This protocol for laboratory-scale production of spider silk, which is stronger than tensile steel, can potentially be extended to large-scale manufacturing.
In JoVE Clinical and Translational Medicine, we featured a number of articles related to stroke, one of which. In one article, M&ouml;bius-Winkler et al. demonstrated placement of the WATCHMAN left atrial appendage occlusion device from Boston Scientific. This device is designed to prevent strokes by trapping clots within the left atrial appendage of the heart before they exit. Our authors demonstrated how to insert the guide wire through the femoral artery into the heart, advance the device into the left atrium, and deploy the device in the left atrial appendage.
Another article focused on stroke, Hamel et al. described the use of a driving simulator to study how patients compensate for visual field defects following stroke. By analyzing the compensatory gaze behavior of patients as they navigated through virtual driving courses with varying degrees of complexity, our authors see great potential for the use of driving simulators in stroke rehabilitation.
In JoVE Immunology and Infection, we published an article by Keyel et al. describing the real-time kinetics of immune cell responses to bacterial toxins using live cell microscopy. Combined with high-speed 3D confocal microscopy, this technique can also visualize the cellular repair response.
Also in JoVE Immunology and Infection, we featured an article by Yap et al. describing methods for diagnosing helminth infections in children. Helminths are parasitic worms that can infect the intestinal tract of humans and animals, and may be distinguished based on their morphology. The same article demonstrated how to measure the impact of helminth infections on physical fitness in children, which is a measure of overall health.
In JoVE Neuroscience, we featured a method by Hoffmann et al., who used tiny headphones to alter the auditory feedback in songbirds, and analyzed the computational and neurophysiological basis of vocal learning in birds when they adjust their singing in response to altered acoustic signals.
Also in JoVE Neuroscience, we moved from birds to bees with a method that assesses associative and non-associative tactile learning in honeybees. This method, by Mujagi&#263; et al., allowed bees to scan different metal surfaces with their antennae, then conditioned them to expect sugar water when their antennae touched particular surfaces, and analyzed the corresponding changes in antennal movement.
In 2012, we launched a brand-new section: JoVE Applied Physics, which features articles on subjects ranging from plasma physics to material science. The Applied Physics section also contained JoVE's 2000th video article, filmed in a synchrotron radiation facility, where Borisenko et al. showed how to determine the electronic structure of complex materials using angle-resolved photoemission spectroscopy.
In the JoVE General section, we published an article by Artioli et al. demonstrating the use of a portable gas analyzer to measure oxygen consumption during complex exercise - such as judo. By analyzing oxygen consumption and collecting blood for measuring plasma lactate concentration, this method can determine the relative contributions of different energy systems to specific aspects of complex exercise.
Finally, JoVE followed Dolhi et al. to a permanently ice-covered saline lake in Antarctica. There, scientists drilled through the ice to study single-celled microorganisms called protists that live in the harsh conditions of Antarctica.
This Year in Review was just a brief glimpse of a few of over 600 video articles that JoVE offered in 2012. Browse the JoVE archives to see thousands of other videos, and stay tuned for what's coming up this year in JoVE: The Journal of Visualized Experiments.

Here's a look at some of the milestones and highlights of the year 2012 in Journal of Visualized Experiments (JoVE).

2012: Das Jahr im Rückblick

 
 
Here's a look at some of the milestones and highlights of the year 2012 in Journal of Visualized Experiments (JoVE).
At the start of 2012, JoVE ...

Dieses November in JoVE

Zusammenfassung

Im Dezember dieses Jahres in JoVE

August 2012: Diesen Monat in JoVE

2012: Das Jahr im Rückblick