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Journal of Visualized Experiments: JoVE

Haesung Park, Thomas W LeBrun
We demonstrate a method to trap a selected dielectric microparticle in air using radiation pressure from a single-beam gradient optical trap. Randomly scattered dielectric microparticles adhered to a glass substrate are momentarily detached using ultrasonic vibrations generated by a piezoelectric transducer (PZT). Then, the optical beam focused on a selected particle lifts it up to the optical trap while the vibrationally excited microparticles fall back to the substrate. A particle may be trapped at the nominal focus of the trapping beam or at a position above the focus (referred to here as the levitation position) where gravity provides the restoring force...
February 5, 2017: Journal of Visualized Experiments: JoVE
Josef Brown, Benjamin F Davis, Matthew R Maschmann
A nanoscale fabrication technique appropriate for milling carbon nanotube (CNT) forests is described. The technique utilizes an environmental scanning electron microscope (ESEM) operating with a low pressure water vapor ambient. In this technique, a portion of the electron beam interacts with the water vapor in the vicinity of the CNT sample, dissociating the water molecules into hydroxyl radicals and other species by radiolysis. The remainder of the electron beam interacts with the CNT forest sample, making it susceptible to oxidation from the chemical products of radiolysis...
February 5, 2017: Journal of Visualized Experiments: JoVE
Justus Hermannsdörfer, Niels de Jonge
Samples fully embedded in liquid can be studied at a nanoscale spatial resolution with Scanning Transmission Electron Microscopy (STEM) using a microfluidic chamber assembled in the specimen holder for Transmission Electron Microscopy (TEM) and STEM. The microfluidic system consists of two silicon microchips supporting thin Silicon Nitride (SiN) membrane windows. This article describes the basic steps of sample loading and data acquisition. Most important of all is to ensure that the liquid compartment is correctly assembled, thus providing a thin liquid layer and a vacuum seal...
February 5, 2017: Journal of Visualized Experiments: JoVE
Irena Roci, Hector Gallart-Ayala, Jeramie Watrous, Mohit Jain, Craig E Wheelock, Roland Nilsson
Mammalian cell types exhibit specialized metabolism, and there is ample evidence that various co-existing cell types engage in metabolic cooperation. Moreover, even cultures of a single cell type may contain cells in distinct metabolic states, such as resting or cycling cells. Methods for measuring metabolic activities of such subpopulations are valuable tools for understanding cellular metabolism. Complex cell populations are most commonly separated using a cell sorter, and subpopulations isolated by this method can be analyzed by metabolomics methods...
February 4, 2017: Journal of Visualized Experiments: JoVE
Holly A Bilton, Zainab Ahmad, Nancy Janzen, Shannon Czorny, John F Valliant
Pre-targeting combined with bioorthogonal chemistry is emerging as an effective way to create new radiopharmaceuticals. Of the methods available, the inverse electron demand Diels-Alder (IEDDA) cycloaddition between a radiolabeled tetrazines and trans-cyclooctene (TCO) linked to a biomolecule has proven to be a highly effective bioorthogonal approach to imaging specific biological targets. Despite the fact that technetium-99m remains the most widely used isotope in diagnostic nuclear medicine, there is a scarcity of methods for preparing (99m)Tc-labeled tetrazines...
February 4, 2017: Journal of Visualized Experiments: JoVE
Kenta Mizuse, Romu Fujimoto, Nobuo Mizutani, Yasuhiro Ohshima
We present a method for visualizing laser-induced, ultrafast molecular rotational wave packet dynamics. We have developed a new 2-dimensional Coulomb explosion imaging setup in which a hitherto-impractical camera angle is realized. In our imaging technique, diatomic molecules are irradiated with a circularly polarized strong laser pulse. The ejected atomic ions are accelerated perpendicularly to the laser propagation. The ions lying in the laser polarization plane are selected through the use of a mechanical slit and imaged with a high-throughput, 2-dimensional detector installed parallel to the polarization plane...
February 4, 2017: Journal of Visualized Experiments: JoVE
Ali Rashidfarrokhi, Veronica Richina, Fikadu G Tafesse
The mammalian body is equipped with various layers of mechanisms that help to defend itself from pathogen invasions. Professional phagocytes of the immune system - such as neutrophils, dendritic cells, and macrophages - retain the innate ability to detect and clear such invading pathogens through phagocytosis(1). Phagocytosis involves choreographed events of membrane reorganization and actin remodeling at the cell surface(2)(,)(3). Phagocytes successfully internalize and eradicate foreign molecules only when all stages of phagocytosis are fulfilled...
February 3, 2017: Journal of Visualized Experiments: JoVE
Altea Rocchi, Congcong He
Autophagy is a lysosomal degradation pathway essential for cell homeostasis, function and differentiation. Under stress conditions, autophagy is induced and targets various cargos, such as bulk cytosol, damaged organelles and misfolded proteins, for degradation in lysosomes. Resulting nutrient molecules are recycled back to the cytosol for new protein synthesis and ATP production. Upregulation of autophagy has beneficial effects against the pathogenesis of many diseases, and pharmacological and physiological strategies to activate autophagy have been reported...
February 3, 2017: Journal of Visualized Experiments: JoVE
Marco Morsch, Rowan A W Radford, Emily K Don, Albert Lee, Elinor Hortle, Nicholas J Cole, Roger S Chung
Using a standard confocal setup, a UV ablation method can be utilized to selectively induce cellular injury and to visualize single-cell responses and cell-cell interactions in the CNS in real-time. Previously, studying these cell-specific responses after injury often required complicated setups or the transfer of cells or animals into different, non-physiological environments, confounding immediate and short-term analysis. For example, drug-mediated ablation approaches often lack the specificity that is required to study single-cell responses and immediate cell-cell interactions...
February 3, 2017: Journal of Visualized Experiments: JoVE
M Angélica Bello, Yolanda Ruiz-León, J Vladimir Sandoval-Sierra, Svetlana Rezinciuc, Javier Diéguez-Uribeondo
Common problems in the processing of biological samples for observations with the scanning electron microscope (SEM) include cell collapse, treatment of samples from wet microenvironments and cell destruction. Using young floral tissues, oomycete cysts, and fungi spores (Agaricales) as examples, specific protocols to process delicate samples are described here that overcome some of the main challenges in sample treatment for image capture under the SEM. Floral meristems fixed with FAA (Formalin-Acetic-Alcohol) and processed with the Critical Point Dryer (CPD) did not display collapsed cellular walls or distorted organs...
February 3, 2017: Journal of Visualized Experiments: JoVE
Wuqiang Zhu, Ling Gao, Jianyi Zhang
Human induced pluripotent stem cells (hiPSCs) must be fully differentiated into specific cell types before administration, but conventional protocols for differentiating hiPSCs into cardiomyocytes (hiPSC-CMs), endothelial cells (hiPSC-ECs), and smooth muscle cells (SMCs) are often limited by low yield, purity, and/or poor phenotypic stability. Here, we present novel protocols for generating hiPSC-CMs, -ECs, and -SMCs that are substantially more efficient than conventional methods, as well as a method for combining cell injection with a cytokine-containing patch created over the site of administration...
February 3, 2017: Journal of Visualized Experiments: JoVE
Amy L Louer, Denise N Simon, Karen M Switkowski, Sheryl L Rifas-Shiman, Matthew W Gillman, Emily Oken
A high proportion of children have overweight and obesity in the United States and other countries. Accurate assessment of anthropometry is essential to understand health effects of child growth and adiposity. Gold standard methods of measuring adiposity, such as dual X-ray absorptiometry (DXA), may not be feasible in large field studies. Research staff can, however, complete anthropometric measurements, such as body circumferences and skinfold measurements, using inexpensive portable equipment. In this protocol we detail how to obtain manual anthropometric measurements from children, including standing and sitting height, weight, waist circumference, hip circumference, mid-upper arm circumference, triceps skinfold thickness, and subscapular skinfold thickness, and procedures to assess the quality of these measurements...
February 2, 2017: Journal of Visualized Experiments: JoVE
Luc Bertrand, Levi Dygert, Michal Toborek
Cerebrovascular disease is highly prevalent in the global population and encompasses several types of conditions, including stroke. To study the impact of stroke on tissue injury and to evaluate the effectiveness of therapeutic interventions, several experimental models in a variety of species were developed. They include complete global cerebral ischemia, incomplete global ischemia, focal cerebral ischemia, and multifocal cerebral ischemia. The model described in this protocol is based on the middle cerebral artery occlusion (MCAO) and is related to the focal ischemia category...
February 2, 2017: Journal of Visualized Experiments: JoVE
Zachary H Gursky, Anna Y Klintsova
Aerobic exercise (e.g., wheel running (WR) extensively used in animal research) positively impacts many measures of neuroplastic potential in the brain, such as rates of adult neurogenesis, angiogenesis, and expression of neurotrophic factors in rodents. This intervention has also been shown to mitigate behavioral and neuroanatomical aspects of the negative impacts of teratogens (i.e., developmental exposure to alcohol) and age-related neurodegeneration in rodents. Environmental complexity (EC) has been shown to produce numerous neuroplastic benefits in cortical and subcortical structures and can be coupled with wheel running to increase the proliferation and survival of new cells in the adult hippocampus...
February 2, 2017: Journal of Visualized Experiments: JoVE
Alexandra Linnemann, Jana Strahler, Urs M Nater
Music listening is associated with stress-reducing effects. However, most of the results on music listening and stress were gathered in experimental settings. As music listening is a popular activity of daily life, it is of utmost importance to study the effects of music listening on psychobiological stress in an everyday, daily-life setting. Here, a study protocol is presented that allows the assessment of associations between music listening and psychobiological stress in daily life by noninvasively measuring salivary cortisol (as a marker of the Hypothalamic-Pituitary-Adrenal (HPA) axis) and salivary alpha-amylase (as a marker of the Autonomic Nervous System (ANS))...
February 2, 2017: Journal of Visualized Experiments: JoVE
Hironori Nishitsuji, Hiromi Yamamoto, Ritsuko Shiina, Keisuke Harada, Saneyuki Ujino, Kunitada Shimotohno
Currently, it is possible to construct recombinant forms of various viruses, such as human immunodeficiency virus 1 (HIV-1) and hepatitis C virus (HCV), that carry foreign genes such as a reporter or marker protein in their genomes. These recombinant viruses usually faithfully mimic the life cycle of the original virus in infected cells and exhibit the same host range dependence. The development of a recombinant virus enables the efficient screening of inhibitors and the identification of specific host factors...
February 1, 2017: Journal of Visualized Experiments: JoVE
Elina Staaf, Sunitha Bagawath-Singh, Sofia Johansson
Fluorescence correlation spectroscopy (FCS) is a powerful technique for studying the diffusion of molecules within biological membranes with high spatial and temporal resolution. FCS can quantify the molecular concentration and diffusion coefficient of fluorescently labeled molecules in the cell membrane. This technique has the ability to explore the molecular diffusion characteristics of molecules in the plasma membrane of immune cells in steady state (i.e., without processes affecting the result during the actual measurement time)...
February 1, 2017: Journal of Visualized Experiments: JoVE
Matthew J Roy, Daan M Maijer
High performance, cast aluminum automotive wheels are increasingly being incrementally formed via flow forming/metal spinning at elevated temperatures to improve material properties. With a wide array of processing parameters which can affect both the shape attained and resulting material properties, this type of processing is notoriously difficult to commission. A simplified, light-duty version of the process has been designed and implemented for full-size automotive wheels. The apparatus is intended to assist in understanding the deformation mechanisms and the material response to this type of processing...
February 1, 2017: Journal of Visualized Experiments: JoVE
Eylon Persky, Anna Kremen, Shai Wissberg, Yishai Shperber, Beena Kalisky
Local, deterministic manipulation of individual vortices in type 2 superconductors is challenging. The ability to control the position of individual vortices is necessary in order to study how vortices interact with each other, with the lattice, and with other magnetic objects. Here, we present a protocol for vortex manipulation in thin superconducting films by local contact, without applying current or magnetic field. Vortices are imaged using a scanning superconducting quantum interference device (SQUID), and vertical stress is applied to the sample by pushing the tip of a silicon chip into the sample, using a piezoelectric element...
February 1, 2017: Journal of Visualized Experiments: JoVE
Ana R Leda, Levy Dygert, Luc Bertrand, Michal Toborek
Animal models of central nervous system (CNS) diseases and, consequently, blood-brain barrier disruption diseases, require the delivery of exogenous substances into the brain. These exogenous substances may induce injurious impact or constitute therapeutic strategy. The most common delivery methods of exogenous substances into the brain are based on systemic deliveries, such as subcutaneous or intravenous routes. Although commonly used, these approaches have several limitations, including low delivery efficacy into the brain...
January 31, 2017: Journal of Visualized Experiments: JoVE
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