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Imaging Cell Adhesion and Migration
Published in Margarida M. Barroso, Xavier Intes, In Vivo, 2020
Chandrani Mondal, Julie Di Martino, Jose Javier Bravo-Cordero
New microscopy techniques that have improved resolution have also been successfully applied to the study of focal adhesions. Super-resolution imaging increases spatial resolution compared to confocal microscopy. The development of techniques such as structured illumination microscopy (SIM), photoactivated localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM), and stimulated emission depletion (STED) enable the characterization of subdomains of focal adhesions and revealed their dynamic architecture with a single-molecule localization scale (Schwartz, 2011; Sahl et al., 2017). All these technologies have contributed to the improvement and understanding of adhesions as well as the establishment of novel models that are shedding light into how cells organize focal adhesions at the molecular level. For example, PALM has revealed that signaling and cytoskeletal proteins reside at specific vertical distances between the plasma membrane and F-actin (Schwartz, 2011).
Single Molecule Fluorescence
Published in Yuri L. Lyubchenko, An Introduction to Single Molecule Biophysics, 2017
A related set of methods that has emerged over the last few years are the so-called super-resolution imaging methods. This is a neat way to get around the classical diffraction limit of imaging, which was the dogma in the imaging field until about a decade back. The idea is that diffraction limits the resolution of two objects below about half the wavelength of the emitted light. However, as noted above, if one images the objects (molecules) one at a time, their positions can be localized with much higher precision (arbitrarily high, depends on the number of collected photons). Hence, groups (initially of Xioawei Zhuang [Rust et al. 2006] and Eric Betzig [Betzig et al. 2006]) have developed methods to stochastically turn on a small fraction of single molecules in a sample, localize them by imaging, turn them off, then turn on another small subset, and so on. Hence, by imaging an ensemble of molecules, one at a time, high-resolution imaging can be performed, which is providing detailed views of cellular architecture that were previously unavailable from fluorescence imaging (Bates et al. 2008; Patterson et al. 2010). Indeed, the impact of this advance is large enough that the 2014 Nobel Prize in Chemistry in part went to Betzig for his above contribution.
Soft X-ray Tomography: Techniques and Applications
Published in Paolo Russo, Handbook of X-ray Imaging, 2017
Axel A. Ekman, Tia E. Plautz, Jian-Hua Chen, Gerry McDermott, Mark A. Le Gros, Carolyn A. Larabell
Ideally, the position of these fluorescent tags in the cell should be determined with the highest possible spatial resolution (Willig et al. 2006). For many years, the spatial resolution was thought to be limited by diffraction, leading to a maximum resolution limit of typically half the wavelength of the light used (Abbe 1873); in other words, 200–300 nm. However, in recent years, a number of “super-resolution” imaging techniques have been developed that extend beyond this resolution (Hell et al. 2004, Betzig et al. 2006, Rust et al. 2006, Bates et al. 2007, Friedenberger et al. 2007, Willig et al. 2007, Manley et al. 2008, Punge et al. 2008, Schermelleh et al. 2008). All of these methods can be used to localize a fluorescent moiety at a spatial resolution higher than the optical diraction limit.
Blind structured illumination as excitation for super-resolution photothermal radiometry
Published in Quantitative InfraRed Thermography Journal, 2020
Peter Burgholzer, Thomas Berer, Mathias Ziegler, Erik Thiel, Samim Ahmadi, Jürgen Gruber, Günther Mayr, Günther Hendorfer
Our method for increasing the spatial resolution was inspired by recent work in fluorescence microscopy, where super-resolution imaging was demonstrated using multiple unknown speckle illumination patterns [8,9]. We extend this concept to thermographic imaging, with several illumination patterns, similar to what we have done for photoacoustic imaging [10]. Figure 1 illustrates one of different illumination patters to illuminate the sample. The illumination patterns and the absorption pattern are represented by discrete vectors , where is the number of acquired camera pixels and the components denote values at equidistant points in the imaging domain. According to Equation (1) the reconstructed thermal signal, measured by the infrared camera is
Light, the universe and everything – 12 Herculean tasks for quantum cowboys and black diamond skiers
Published in Journal of Modern Optics, 2018
Girish Agarwal, Roland E. Allen, Iva Bezděková, Robert W. Boyd, Goong Chen, Ronald Hanson, Dean L. Hawthorne, Philip Hemmer, Moochan B. Kim, Olga Kocharovskaya, David M. Lee, Sebastian K. Lidström, Suzy Lidström, Harald Losert, Helmut Maier, John W. Neuberger, Miles J. Padgett, Mark Raizen, Surjeet Rajendran, Ernst Rasel, Wolfgang P. Schleich, Marlan O. Scully, Gavriil Shchedrin, Gennady Shvets, Alexei V. Sokolov, Anatoly Svidzinsky, Ronald L. Walsworth, Rainer Weiss, Frank Wilczek, Alan E. Willner, Eli Yablonovitch, Nikolay Zheludev
Optical superoscillations were observed experimentally by the Southampton group in 2007 [120]. In a superoscillatory focus, only a small fraction of energy goes into the hotspot, while the majority of the light’s energy is distributed into a broad “halo” surrounding the hotspot. The area between the hotspot and the surrounding halo is often referred to as the “field of view”. Computer modelling shows that objects smaller than the field of view can be imaged in a scanning mode with resolution corresponding to the size of the hot spot. For instance a pair of 9 nm x 9 nm holes in an opaque screen, spaced by 28 nm, can be perfectly resolved by a superoscillatory imaging apparatus operating at the wavelength of 400 nm, thus giving resolution exceeding one-tenth of the wavelength [214]. In the last few years, a practical label-free super-resolution imaging technology based on superoscillations combined with confocal detection has been developed and resolution up to one six of the wavelength has already been demonstrated [215]. This technique is now deployed in biological imaging [216].