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https://www.readbyqxmd.com/read/28943243/hucmsc-exosome-transported-14-3-3%C3%AE-prevents-the-injury-of-cisplatin-to-hk-2-cells-by-inducing-autophagy-in-vitro
#1
REVIEW
Juanjuan Wang, Haoyuan Jia, Bin Zhang, Lei Yin, Fei Mao, Jing Yu, Cheng Ji, Xiao Xu, Yongmin Yan, Wenrong Xu, Hui Qian
BACKGROUND AIMS: On the basis of previous studies, exosomes secreted by human umbilical cord mesenchymal stromal cell (hucMSC-ex) could prevent and repair acute kidney injury induced by cisplatin in rats. However, its potential mechanism is still unclear. In the present study, the model with hucMSC-ex pretreated human renal tubular epithelial cell lines HK-2 that could prevent the injury of cisplatin was successfully established. METHODS: First, we pretreated the HK-2 cells with hucMSC-ex for 24 h...
September 21, 2017: Cytotherapy
https://www.readbyqxmd.com/read/28940069/purification-and-application-of-a-small-actin-probe-for-single-molecule-localization-microscopy
#2
Roderick P Tas, Trusanne G A A Bos, Lukas C Kapitein
The cytoskeleton is involved in many cellular processes. Over the last decade, super-resolution microscopy has become widely available to image cytoskeletal structures, such as microtubules and actin, with great detail. For example, Single-Molecule Localization Microscopy (SMLM) achieves resolutions of 5-50 nm through repetitive sparse labeling of samples, followed by Point-Spread-Function analysis of individual fluorophores. Whereas initially this approach depended on the controlled photoswitching of fluorophores targeted to the structure of interest, alternative techniques now depend on the transient binding of fluorescently labeled probes, such as the small polypeptide lifeAct that can transiently interact with polymerized actin...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28940065/a-brief-introduction-to-single-molecule-fluorescence-methods
#3
Siet M J L van den Wildenberg, Bram Prevo, Erwin J G Peterman
One of the more popular single-molecule approaches in biological science is single-molecule fluorescence microscopy, which will be the subject of the following section of this volume. Fluorescence methods provide the sensitivity required to study biology on the single-molecule level, but they also allow access to useful measurable parameters on time and length scales relevant for the biomolecular world. Before several detailed experimental approaches will be addressed, we will first give a general overview of single-molecule fluorescence microscopy...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28939861/multiplexed-exchange-paint-imaging-reveals-ligand-dependent-egfr-and-met-interactions-in-the-plasma-membrane
#4
Jeffrey L Werbin, Maier S Avendaño, Verena Becker, Ralf Jungmann, Peng Yin, Gaudenz Danuser, Peter K Sorger
Signal transduction by receptor tyrosine kinases (RTKs) involves complex ligand- and time-dependent changes in conformation and modification state. High resolution structures are available for individual receptors dimers, but less is known about receptor clusters that form in plasma membranes composed of many different RTKs with the potential to interact. We report the use of multiplexed super-resolution imaging (Exchange-PAINT) followed by mean-shift clustering and random forest analysis to measure the precise distributions of five receptor tyrosine kinases (RTKs) from the ErbB, IGF-1R and Met families in breast cancer cells...
September 22, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28935861/correlation-profiling-of-brain-sub-cellular-proteomes-reveals-co-assembly-of-synaptic-proteins-and-subcellular-distribution
#5
Nikhil J Pandya, Frank Koopmans, Johan A Slotman, Iryna Paliukhovich, Adriaan B Houtsmuller, August B Smit, Ka Wan Li
Protein correlation profiling might assist in defining co-assembled proteins and subcellular distribution. Here, we quantified the proteomes of five biochemically isolated mouse brain cellular sub-fractions, with emphasis on synaptic compartments, from three brain regions, hippocampus, cortex and cerebellum. We demonstrated the expected co-fractionation of canonical synaptic proteins belonging to the same functional groups. The enrichment profiles also suggested the presence of many novel pre- and post-synaptic proteins...
September 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28933862/creim-coffee-ring-effect-imaging-model-for-monitoring-protein-self-assembly-in-situ
#6
Michael Shaw, Angelo Bella, Maxim G Ryadnov
Protein self-assembly is fundamental to nanotechnology. Self-assembling structures are produced under static in vitro conditions typically forming over hours. In contrast, hydrodynamic intracellular environments employ far shorter time scales to compartmentalize highly concentrated protein solutions. Herein, we exploit the radial capillary flow within a drying sessile droplet (the coffee ring effect) to emulate dynamic native environments and monitor an archetypal protein assembly in situ using high-speed super-resolution imaging...
September 22, 2017: Journal of Physical Chemistry Letters
https://www.readbyqxmd.com/read/28930989/demonstration-of-a-hyperlens-integrated-microscope-and-super-resolution-imaging
#7
Dasol Lee, Minkyung Kim, Sunae So, Inki Kim, Gwanho Yoon, Kyunghoon Kim, Junsuk Rho
The use of super-resolution imaging to overcome the diffraction limit of conventional microscopy has attracted the interest of researchers in biology and nanotechnology. Although near-field scanning microscopy and superlenses have improved the resolution in the near-field region, far-field imaging in real-time remains a significant challenge. Recently, the hyperlens, which magnifies and converts evanescent waves into propagating waves, has emerged as a novel approach to far-field imaging. Here, we report the fabrication of a spherical hyperlens composed of alternating silver (Ag) and titanium oxide (TiO2) thin layers...
September 8, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28926050/temperature-dependence-of-the-conversion-efficiency-of-photochromic-perylene-bisimide-dithienylcyclopentene-triads-embedded-in-a-polymer
#8
Johann Thurn, Johannes Maier, Martti Pärs, Katja Gräf, Mukundan Thelakkat, Jürgen Köhler
Photochromic molecules that are covalently linked to a strong fluorophore combine the requirements of external control and strong fluorescence, which will become increasingly important for super-resolution microscopy techniques based on single molecules. However, given the bulky structure of such constructs, steric hindrance might affect their photoconversion efficiencies upon immobilising them for imaging purposes. In this study the efficiencies of the photochromic conversion processes of molecular triads that are embedded in a polymer have been studied as a function of temperature...
September 19, 2017: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/28924673/super-resolution-high-content-screening-and-analysis
#9
K Soliman
Revealing the subcellular phenotypes at the molecular scale is critical to understand the mechanisms by which the cells function and respond to chemical treatments. Super-resolution microscopy and robust analysis tools enabled biologists to reveal and quantify phenotypes at unprecedented resolution. Developing automated imaging analysis solutions for super-resolution imaging will make high-content-screening (HCS) applicable for super-resolution microscopy, which will give access to new complex information. Here, I provide an instant automated analysis procedure for analyzing super-resolution images via CellProfiler ( www...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924672/in-situ-super-resolution-imaging-of-genomic-dna-with-oligostorm-and-oligodna-paint
#10
Brian J Beliveau, Alistair N Boettiger, Guy Nir, Bogdan Bintu, Peng Yin, Xiaowei Zhuang, C-Ting Wu
OligoSTORM and OligoDNA-PAINT meld the Oligopaint technology for fluorescent in situ hybridization (FISH) with, respectively, Stochastic Optical Reconstruction Microscopy (STORM) and DNA-based Point Accumulation for Imaging in Nanoscale Topography (DNA-PAINT) to enable in situ single-molecule super-resolution imaging of nucleic acids. Both strategies enable ≤20 nm resolution and are appropriate for imaging nanoscale features of the genomes of a wide range of species, including human, mouse, and fruit fly (Drosophila)...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924671/correlative-fluorescence-super-resolution-localization-microscopy-and-platinum-replica-em-on-unroofed-cells
#11
Kem A Sochacki, Justin W Taraska
Platinum replicas of unroofed mammalian cells can be imaged with a transmission electron microscope (TEM) to produce high contrast, high resolution images of the structure of the cytoplasmic side of a plasma membrane. A complementary approach, super-resolution fluorescence localization microscopy, can be used to localize labeled molecules with better than 20 nm precision in cells. Here, we describe a correlative method that couples these two techniques and produces images where localization microscopy data can be used to highlight specific proteins of interest within the structural context of the platinum replica TEM image...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924669/measuring-nanometer-distances-between-fluorescent-labels-step-by-step
#12
Susanna Maria Früh, Ingmar Schoen
Super-resolution fluorescence microscopy methods are increasingly applied to study the structure of biological molecules within their natural context or at biomaterial interfaces. We here provide a protocol for Single-molecule High-Resolution Imaging with Photobleaching (SHRImP) that can be used to obtain information about the conformation of large proteins or other macromolecules at the single-molecule level. This procedure requires site-specific protein labeling with fluorescent dyes, immobilization and sample preparation, optimization of imaging buffer composition and microscope settings, and acquisition of short time-lapse movies that capture the stepwise bleaching behavior of individual molecules...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924667/correlative-in-resin-super-resolution-fluorescence-and-electron-microscopy-of-cultured-cells
#13
Errin Johnson, Rainer Kaufmann
Correlative super-resolution light and electron microscopy (super-resolution CLEM) is a powerful and emerging tool in biological research. The practical realization of these two very different microscopy techniques with their individual requirements remains a challenging task. There is a broad range of approaches to choose from, each with their own advantages and limitations. Here, we present a detailed protocol for in-resin super-resolution CLEM of high-pressure frozen and freeze substituted cultured cells...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924666/brain-slice-staining-and-preparation-for-three-dimensional-super-resolution-microscopy
#14
Christopher L German, Manasa V Gudheti, Annette E Fleckenstein, Erik M Jorgensen
Localization microscopy techniques-such as photoactivation localization microscopy (PALM), fluorescent PALM (FPALM), ground state depletion (GSD), and stochastic optical reconstruction microscopy (STORM)-provide the highest precision for single-molecule localization currently available. However, localization microscopy has been largely limited to cell cultures due to the difficulties that arise in imaging thicker tissue sections. Sample fixation and antibody staining, background fluorescence, fluorophore photoinstability, light scattering in thick sections, and sample movement create significant challenges for imaging intact tissue...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924663/quantitative-single-molecule-localization-microscopy-qsmlm-of-membrane-proteins-based-on-kinetic-analysis-of-fluorophore-blinking-cycles
#15
Franziska Fricke, Joel Beaudouin, Sebastian Malkusch, Roland Eils, Mike Heilemann
Photoswitchable or photoactivatable fluorophores are the key in single-molecule localization microscopy. Next to providing fluorescence images with subdiffraction spatial resolution, additional information is available from observing single fluorophores over time. This includes the characteristic photophysical phenomenon of "blinking" that is exhibited by single fluorescent proteins or fluorophores and follows well-defined kinetic laws. Analyzing the kinetics of "blinking" allows determining the number of fluorophores in a multi-molecular complex...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924662/correlative-super-resolution-fluorescence-imaging-and-atomic-force-microscopy-for-the-characterization-of-biological-samples
#16
Patricia Bondia, Santiago Casado, Cristina Flors
Recent advances in imaging tools have greatly improved our ability to analyze the structure and molecular components of a wide range of biological systems at the nanoscale. High resolution imaging can be performed with a handful of techniques, each of them revealing particular features of the sample. A more comprehensive picture of a biological system can be achieved by combining the information provided by complementary imaging methods. Specifically, the correlation between super-resolution fluorescence imaging and atomic force microscopy (AFM) provides high resolution topography as well as specific chemical information, the latter with a spatial resolution that approaches that of AFM...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924660/four-channel-super-resolution-imaging-by-3-d-structured-illumination
#17
Ulrike Engel
Multichannel imaging is used as a readout of relative localization of two or more components and is often the first step in investigating functional ensembles in cells. However, the localization volume of diffraction-limited light microscopy (approx. 200 nm by 500 nm) can accommodate hundred of proteins, calling for increased resolution for these types of analyses. Here, we present a protocol for 4-channel imaging using structured illumination microscopy (SIM), which increases resolution by a factor of two...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924659/sted-imaging-of-golgi-dynamics-with-cer-sir-a-two-component-photostable-high-density-lipid-probe-for-live-cells
#18
Roman S Erdmann, Derek Toomre, Alanna Schepartz
Long time-lapse super-resolution imaging in live cells requires a labeling strategy that combines a bright, photostable fluorophore with a high-density localization probe. Lipids are ideal high-density localization probes, as they are >100 times more abundant than most membrane-bound proteins and simultaneously demark the boundaries of cellular organelles. Here, we describe Cer-SiR, a two-component, high-density lipid probe that is exceptionally photostable. Cer-SiR is generated in cells via a bioorthogonal reaction of two components: a ceramide lipid tagged with trans-cyclooctene (Cer-TCO) and a reactive, photostable Si-rhodamine dye (SiR-Tz)...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924658/two-photon-sted-microscopy-for-nanoscale-imaging-of-neural-morphology-in-vivo
#19
Mirelle J T Ter Veer, Thomas Pfeiffer, U Valentin Nägerl
The advent of super-resolution microscopy offers to bridge the gap between electron and light microscopy. It has opened up the possibility of visualizing cellular structures and dynamic signaling events on the "mesoscale" well below the classic diffraction barrier of light microscopy (10-200 nm), while essentially retaining the advantages of fluorescence microscopy concerning multicolor labeling, detection sensitivity, signal contrast, live-cell imaging, and temporal resolution.From among the new super-resolution techniques, STED microscopy stands out as a laser-scanning imaging modality, which enables nanoscale volume-metric imaging of cellular morphology...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28924656/live-cell-sted-imaging-with-the-hyper2-biosensor
#20
Natalia M Mishina, Vsevolod V Belousov
Stimulated emission depletion (STED) microscopy is a popular super resolution imaging technique. Not only synthetic dyes and fluorescent proteins can be utilized as STED fluorophores, but also genetically encoded biosensors. Fusing the biosensor with proteins of interest allows subdiffraction imaging of intracellular macromolecular architecture with simultaneous extraction of functional information about cellular activities. Here, we describe a protocol for live-cell STED microscopy of the HyPer2 biosensor fused to cytoskeletal filaments...
2017: Methods in Molecular Biology
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