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  • mTORC1 Promotes Proliferation of Immature Schwann Cells and Myelin Growth of Differentiated Schwann Cells

    The myelination of axons is essential for neuronal wiring and normal nervous system functions. In the peripheral nervous system, Schwann cells (SCs) form myelin sheaths around axons during nerve development. Such myelination is compromised in a number of diseases. Hence, identification and understanding of the key pathways regulating SC development and myelinogenesis are essential for therapeutic progress. Here we uncover two separate roles of the cellular signaling node mTORC1 (mechanistic target of rapamycin complex 1) for regulating the development of SCs and subsequently the growth of myelin sheaths. Moreover, we demonstrate that defective SCs possess a remarkable plasticity to remyelinate axons via mTORC1. Thus, manipulating mTORC1 activity in diseased SCs could be therapeutically beneficial.
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  • FCS - Fluorescence Correlation Spectroscopy

    FCS is a fluorescence-based measurement method. Fluorescent molecules passing through a strongly focused, fixed laser beam are excited for fluorescence emission. After passing a confocal pinhole, the emitted photons are registered using very sensitive detectors.
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  • FLIM FRET - Fluorescence Resonance Energy Transfer

    A typical application of FLIM is FLIM-FRET. FRET is a well-established technique to study molecular interactions. It scrutinizes protein binding and estimates intermolecular distances on an Angström scale as well. The SP8 FALCON system together with the integrated FRET analyzer provides FRET-efficiency and binding maps.
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  • FLCS - Fluorescence Lifetime Correlation Spectroscopy

    Essentially, FCS can be performed with a continuous-wave laser. Using pulsed lasers allows even more sophisticated analysis possibilities, such as time-gated FCS or Fluorescence Lifetime Correlation Spectroscopy (FLCS). Both methods make use of the additional information obtained by the simultaneous measurement of the fluorescence lifetime.
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  • FLIM - Fluorescence Lifetime Imaging

    The fluorescence lifetime is a measure of how long a fluorophore remains on average in its excited state before returning to the ground state by emitting a fluorescence photon.
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  • SP FLIM - Spectral Fluorescence Lifetime Imaging

    The SP8 FALCON is the ideal tool for spectral FLIM detection. No emission filters in front of the FLIM detectors are necessary. This grants a much higher flexibility to the experimental design.
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  • FCCS - Fluorescence Cross-Correlation Spectroscopy

    FCCS (Fluorescence Cross-Correlation Spectroscopy) can be measured using the Leica TCS SP8 FCS system. Similar to FCS , it analyzes fluorescence intensity fluctuations derived from a small observation volume.
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  • IT Security Whitepaper

    Security is a primary concern of Leica Microsystems and Leica Microsystems’ customers that employ remote services. Leica Microsystems requires a proven remote service solution that protects against viruses and hackers and supports our intelligent instruments without major end-user modifications, while working within our current network security model and achieving official certification by a third-party security company.
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  • Expression Analysis of Platelet‐derived Growth Factor Receptor Alpha and its Ligands in the Developing Mouse Lung

    Activation of the platelet‐derived growth factor receptor‐α (PDGFR α) signaling pathway is critically important during lung alveogenesis, the process in lung development during which alveoli are formed from the terminal alveolar sacs. Several studies have aimed to characterize the expression patterns of PDGFR α and its two ligands (PDGF‐A and ‐C) in the lung, but published analyses have been limited to embryonic and/or perinatal time points, and no attempts have been made to characterize both receptor and ligand expression simultaneously. In this study, we present a detailed map of the expression patterns of PDGFR α, PDGF‐A and PDGF‐C during the entire period of lung development, that is, from early embryogenesis until adulthood.
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  • STED image of peripheral section of HeLa cell nucleus

    Abstracts of the 7th European Super-Resolution User-Club Meeting

    The 7th Super-Resolution User Club Meeting was held in collaboration with Prof Pavel Hozák , at the Institute of Molecular Genetics of the ASCR in Prague. Keeping the event close to science is one of the founding principles of the event, allowing all participants to network, share and explore exciting new super-resolution and nanoscopy applications. Central to this are the scientific talks given during the meeting, with this cutting-edge microscopy technique as their central theme. A wide selection of topics were covered, prompting interesting discussions during the workshops.
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  • Primary Beam Splitting Devices for Confocal Microscopes

    Current fluorescence microscopy employs incident illumination which requires separation of illumination and emission light. The classical device performing this separation is a color-dependent beam splitting mirror which has fixed spectral parameters and transmits the emission usually between 90% and 98% within the designated bands. Transmission is wavelength dependent and also differs by technology, requirements and design. An alternative is the acousto optical beam splitter which has freely tunable reflection notches and transmits the emission on average at 95% between these notches.
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  • Pinhole Effect in Confocal Microscopes

    When operating a confocal microscope, or when discussing features and parameters of such a device, we inescapably mention the pinhole and its diameter. This short introductory document is meant to explain the significance of the pinhole for those, who did not want to spend too much time to dig into theory and details of confocal microscopy but wanted to have an idea about the effect of the pinhole.
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  • Clarifying Tissue Clearing

    Biological specimens are intrinsically three dimensional; however because of the obscuring effects of light scatter, imaging deep into a tissue volume is problematic. Although efforts to eliminate the scatter by “clearing” the tissue have been ongoing for over a century, there have been a large number of recent innovations. This review introduces the physical basis for light-scatter in tissue, describes the mechanisms underlying various clearing techniques, and discusses several of the major advances in light microscopy for imaging cleared tissue.
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  • Chronic Inflammation Under the Microscope

    In the course of chronic inflammation certain body areas are recurrently inflamed. This goes along with many human diseases. With the help of widefield light microscopy, the underlying processes can be examined from a cellular level to whole organisms. This article presents several widefield microscopy applications such as immunofluorescence, live-cell imaging, histology, and ratiometric analysis to get insight into the development of chronic inflammation, the related diseases, and their treatment.
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  • Methods to Calibrate and Scale Axial Distances in Confocal Microscopy as a Function of Refractive Index

    Application example of HyVolution Super-Resolution - Accurate distance measurement in 3D confocal microscopy is important for quantitative analysis, volume visualization and image restoration. However, axial distances can be distorted by both the point spread function (PSF) and by a refractive-index mismatch between the sample and immersion liquid, which are difficult to separate. Additionally, accurate calibration of the axial distances in confocal microscopy remains cumbersome, although several high-end methods exist. In this paper we present two methods to calibrate axial distances in 3D confocal microscopy that are both accurate and easily implemented.
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  • Multispectral Phloem-Mobile Probes: Properties and Applications

    Using Arabidopsis (Arabidopsis thaliana) seedlings, we identified a range of small fluorescent probes that entered the translocation stream and were unloaded at the root tip. These probes had absorbance/emission maxima ranging from 367/454 to 546/576 nm and represent a versatile toolbox for studying phloem transport. Of the probes that we tested, naturally occurring fluorescent coumarin glucosides (esculin and fraxin) were phloem loaded and transported in oocytes by the sucrose transporter, AtSUC2. Arabidopsis plants in which AtSUC2 was replaced with barley (Hordeum vulgare) sucrose transporter (HvSUT1), which does not transport esculin in oocytes, failed to load esculin into the phloem.
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  • P53- and Mevalonate Pathway–Driven Malignancies Require Arf6 for Metastasis and Drug Resistance

    Application example of HvYolution Super-Resolution - Drug resistance, metastasis, and a mesenchymal transcriptional program are central features of aggressive breast tumors. The GTPase Arf6, often overexpressed in tumors, is critical to promote epithelial–mesenchymal transition and invasiveness. The metabolic mevalonate pathway (MVP) is associated with tumor invasiveness and known to prenylate proteins, but which prenylated proteins are critical for MVP-driven cancers is unknown. We show here that MVP requires the Arf6-dependent mesenchymal program.
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  • Adeno-associated Viral Vectors do not Efficiently Target Muscle Satellite Cells

    Adeno-associated viral (AAV) vectors are becoming an important tool for gene therapy of numerous genetic and other disorders. Several recombinant AAV vectors (rAAV) have the ability to transduce striated muscles in a variety of animals following intramuscular and intravascular administration, and have attracted widespread interest for therapy of muscle disorders such as the muscular dystrophies. Here we examined the relative ability of rAAV vectors derived from AAV6 to target myoblasts, myocytes, and myotubes in culture and satellite cells and myofibers in vivo. AAV vectors are able to transduce proliferating myoblasts in culture, albeit with reduced efficiency relative to postmitotic myocytes and myotubes. In contrast, quiescent satellite cells are refractory to transduction in adult mice.
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  • HyVolution – Super-Resolution Imaging with a Confocal Microscope

    Since the invention of the microscope, there has been continual discussion about the possibility of showing more detailed features of specimens as compared to just magnifying them. In this article we describe the HyVolution concept and how the combination of confocal multiparameter fluorescence imaging at the confocal super-resolution regime with psf-based real deconvolution allows high-speed multicolor imaging with a resolution down to 140 nm.
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  • HyVolution – the Smart Path to Confocal Super-Resolution

    Super-resolution refers to any device or method that can resolve better than the classical Abbe limit. Apart from infinite super-resolution techniques such as STED (stimulated emission depletion) and SMLM (single-molecule localization methods) that can theoretically resolve to any detail, there are also methods for limited super-resolution. Here we present HyVolution by Leica, which merges optical super-resolution and computational super-resolution. The optical part is provided by confocal microscopy, and the computational part by deconvolution. Lateral resolution of 140 nm is demonstrated. HyVolution offers multiple fluorescence recording in truly simultaneous mode.
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  • FRAP with TCS SP8 Resonant Scanner

    Fast FRAP experiments need a sufficient number of measurement points for meaningful interpretation and fitting analysis. To study very fast translocational processes, the use of a resonant scanner (RS) is preferred. The advantage in using FRAP with the RS is that statistics are much better in experiments that require fast acquisition: If the half time of recovery is about 0.5 sec you may have only about 3 to 4 data points using the conventional scanner, whereas with the resonant scanner you can get about 20 data points.
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  • Light Sheet Microscopy Turned Vertically

    Living cells and organisms often suffer from the high light intensities used for fluorescent imaging. Light sheet microscopy reduces phototoxic effects and bleaching by illuminating a specimen in only a single plane at a time. A new light sheet microscope combines light sheet and confocal microscopy in one system without compromising either functionality and allows the combination of the two methods, e.g. confocal photomanipulation with subsequent light sheet acquisition, for new applications.
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  • Progressive Glucose Stimulation of Islet Beta Cells Reveals a Transition From Segregated to Integrated Modular Functional Connectivity Patterns

    Collective beta cell activity in islets of Langerhans is critical for the supply of insulin within an organism. In order to get a detailed insight into the functional organization of the syncytium, we applied advanced analytical tools from the realm of complex network theory to uncover the functional connectivity pattern among cells composing the intact islet.
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  • From Light to Mind: Sensors and Measuring Techniques in Confocal Microscopy

    This article outlines the most important sensors used in confocal microscopy. By confocal microscopy, we mean "True Confocal Scanning", i.e. the technique that illuminates and measures one single point only. The aim is not to impart in-depth specialist knowledge, but to give the user a small but clear overview of the differences between the various technologies and to advise on which sensor may be most suitable for which applications.
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  • "We can go home and the imaging is done automatically by the Leica HCS A Matrix Screener."

    Jutta Maria Bulkescher is the technical coordinator in the Novo Nordisk Foundation Center for Protein Research and Danish Stem Cell Center in Copenhagen, Denmark. The Leica HCS-A matrix screener is an invaluable tool for her facility. "It just gives us the biggest and easiest flexibility we can have to set up different imaging paramters and to check different conditions on one multi-well plate", explains Bulkescher.
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  • Detailed Morphological Characterisation of Hendra Virus Infection of Different Cell Types Using Super-Resolution and Conventional Imaging

    Hendra virus (HeV) is a pleomorphic virus belonging to the Paramyxovirus family. Our long-term aim is to understand the process of assembly of HeV virions. As a first step, we sought to determine the most appropriate cell culture system with which to study this process, and then to use this model to define the morphology of the virus and identify the site of assembly by imaging key virus encoded proteins in infected cells.
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  • Confocal and Digital Light Sheet Imaging

    Optical imaging instrumentation can magnify tiny objects, zoom in on distant stars and reveal details that are invisible to the naked eye. But it notoriously suffers from an annoying problem: the limited depth of field. Our eye-lens (an optical imaging instrument) has the same trouble, but our brain smartly removes all not-in-focus information before the signal reaches conscious cognition.
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  • Venturing into Uncharted Dimensions – the Fascination of Future Technologies

    Beyond the confines of the school curriculum, the young talent promotion program Initiative Junge Forscherinnen und Forscher e.V. (Initiative for Young Researchers) from Würzburg, Bavaria, has set itself the task of fostering an enthusiasm for natural sciences and future technologies in young people. Christoph Stolzenberger is one of the IJF’s science presenters. In his Experimentarium and NanoShuttle, he and his team of postgraduates inspire young researchers’ interest in the wonderful world of microstructures.
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  • Video: High Speed Scanning – With two Scanners in one System

    High speed scanning is necessary to image rapidly changing biological processes. With traditional scanning techniques, imaging speed is limited by the number of fluorophores in a specimen. And, rapid acquisition often comes at the cost of image resolution.
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