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Signal-to-Noise

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  • Webinar: STED Nanoscopy in Combination with Optical Clearing Reveals Localization of Slit Diaphragm Proteins in the Kidney

    In this webinar, we show that optical clearing drastically increases the signal-to-noise ratio and staining quality, thus enabling STED nanoscopy of the subtlest elements of the kidney. In this way we show that optical clearing is not only a sample preparation technique to consider when imaging large mm-scale samples, but could also be fruitful when imaging at the nanoscale. Furthermore, the increased transparency of the optically cleared sample enables volumetric 3D STED imaging at sub-diffraction-limited resolution.
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  • Definitions of Basic Technical Terms for Digital Microscope Cameras and Image Analysis

    Most microscopes today are operated with a camera. The characteristics of the camera often decide whether the acquired image will reveal what a researcher wants to see. But when diving into camera terminology, the technical terms can be overwhelming. We have compiled the most important terms with a concise explanation to provide orientation.
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  • What Makes sCMOS Microscope Cameras so Popular?

    sCMOS cameras are more sensitive and are capable of much higher acquisition speed than cameras with other sensor types. Even though CCD cameras are widely used in live cell imaging and time-lapse recordings, researchers are often concerned that their camera does not detect faint signals. In this interview, Dr. Karin Schwab, Product Manager at Leica Microsystems, talks about the characteristics of sCMOS cameras and how researchers benefit from the latest camera sensor technology.
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  • Introduction to Digital Camera Technology

    A significant majority of modern optical microscopy techniques require the use of a digital camera. By working with digital devices researchers can observe specimens on a screen in real time or acquire and store images and quantifiable data. Here we introduce the basic principles behind digital camera technologies commonly encountered in scientific imaging.
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  • Gated STED Microscopy with Time-gated Single-photon Avalanche Diode

    The maximization of the useful (within the time gate) photon flux is then an important aspect to obtain super-resolved STED images. Here we show that by using a fast-gated single-photon avalanche diode (SPAD), i.e. a detector able to rapidly (hundreds picoseconds) switch-on and -off can improve significantly the signal-to-noise ratio (SNR) of the gated STED image. In addition to an enhancement of the image SNR, the use of the fast-gated SPAD reduces also the system complexity. We demonstrate these abilities both on calibration and biological sample.
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  • Video Talk by Daniel Axelrod: Total Internal Reflection Fluorescence (TIRF) Microscopy

    Total Internal Reflection Fluorescence (TIRF) Microscopy is a technique that only illuminates dye molecules near a surface. In this video, the pioneer of TIRF Microscopy describes what this technique is used for, explains the principles of the evanescent wave, gives many examples of different microscope configurations used in TIRF, and shows how polarized light TIRF can be used to image membrane orientation.
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  • Multi-Images Deconvolution Improves Signal-to-Noise Ratio on Gated Stimulated Emission Depletion Microscopy

    Time-gated detection, namely, only collecting the fluorescence photons after a time-delay from the excitation events, reduces complexity, cost, and illumination intensity of a stimulated emission depletion (STED) microscope. In the gated continuous-wave- (CW-) STED implementation, the spatial resolution improves with increased time-delay, but the signal-to-noise ratio (SNR) reduces.
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  • Huygens STED Deconvolution Quick Guide

    This document is intended to give Leica STED users a brief introduction to deconvolving images on Huygens Professional using images acquired with the Leica TCS SP8 STED 3X microscope. For a more detailed description of Huygens Professional, including additional features and tools, please visit the Manuals section of SVI (www.svi.nl).
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  • Stereo microscopes with TripleBeam Technology

    Especially in fluorescence microscopy, excitation light is friend and foe in one. On the one hand, energy-rich excitation via a specific light wavelength of the fluorochrome resulting in a bright positive fluorochrome signal is highly welcome. On the other hand, "noise" caused by reflections of excitation light passing through the surfaces of optical elements needs to be extremely slight to generate a perfect black background. This relation is described as "signal-to-noise ratio", which is highly relevant for differentiating optically between fluorescence positive and negative cells.
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  • Sensors for True Confocal Scanning

    In this article, advantages and disadvantages of different types of sensors for single point true confocal scanning devices are discussed. Traditionally, photomultiplier tubes have been employed in such systems. For some cases, avalanche photodiodes have proven to fit best. A new development uniting vacuum and silicon technology has led to chimeric sensors, called hybrid detectors (HyD). They benefit from both technologies.
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  • Controlling the TIRF Penetration Depth is Mandatory for Reproducible Results

    The main feature of total internal reflection fluorescence (TIRF) microscopy is the employment of an evanescent wave for the excitation of fluorophores instead of using direct light. A property of the evanescent wave, which arises from the glass/water or glass/specimen interface, is that its propagation in z-direction gradually degrades, limiting its penetration depth into the specimen to some hundred nanometers.
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  • Total Internal Reflection Fluorescence (TIRF) Microscopy

    Total internal reflection fluorescence (TIRF) is a special technique in fluorescence microscopy developed by Daniel Axelrod at the University of Michigan, Ann Arbor in the early 1980s. TIRF microscopy delivers images with an outstandingly high axial resolution below 100 nm. This allows the observation of membrane-associated processes.
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  • Applications of TIRF Microscopy in Life Science Research

    The special feature of TIRF microscopy is the employment of an evanescent field for fluorophore excitation. Unlike standard widefield fluorescence illumination procedures with arc lamps, LEDs or lasers, the evanescent field only penetrates the specimen by about 100 nm starting from the coverslip/medium interface.
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  • A mTurquoise-Based cAMP Sensor for Both FLIM and Ratiometric Read-Out Has Improved Dynamic Range

    FRET-based sensors for cyclic Adenosine Mono Phosphate (cAMP) have revolutionized the way in which this important intracellular messenger is studied. The currently prevailing sensors consist of the cAMP-binding protein Epac1, sandwiched between suitable donor- and acceptor fluorescent proteins (FPs).
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