Zeitschriftenartikel: „Cell microscopy“

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Radosavljević, Jasna Simonović, Aleksandra Lj Mitrović, Ksenija Radotić, László Zimányi, Győző Garab und Gábor Steinbach. „Differential Polarization Imaging of Plant Cells. Mapping the Anisotropy of Cell Walls and Chloroplasts“. International Journal of Molecular Sciences 22, Nr. 14 (17.07.2021): 7661. http://dx.doi.org/10.3390/ijms22147661.

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Modern light microscopy imaging techniques have substantially advanced our knowledge about the ultrastructure of plant cells and their organelles. Laser-scanning microscopy and digital light microscopy imaging techniques, in general—in addition to their high sensitivity, fast data acquisition, and great versatility of 2D–4D image analyses—also opened the technical possibilities to combine microscopy imaging with spectroscopic measurements. In this review, we focus our attention on differential polarization (DP) imaging techniques and on their applications on plant cell walls and chloroplasts, and show how these techniques provided unique and quantitative information on the anisotropic molecular organization of plant cell constituents: (i) We briefly describe how laser-scanning microscopes (LSMs) and the enhanced-resolution Re-scan Confocal Microscope (RCM of Confocal.nl Ltd. Amsterdam, Netherlands) can be equipped with DP attachments—making them capable of measuring different polarization spectroscopy parameters, parallel with the ‘conventional’ intensity imaging. (ii) We show examples of different faces of the strong anisotropic molecular organization of chloroplast thylakoid membranes. (iii) We illustrate the use of DP imaging of cell walls from a variety of wood samples and demonstrate the use of quantitative analysis. (iv) Finally, we outline the perspectives of further technical developments of micro-spectropolarimetry imaging and its use in plant cell studies.

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Yang, Shuntao. „Digital holographic microscopy of highly sensitive living cells“. Journal of Computational Methods in Sciences and Engineering 21, Nr. 6 (07.12.2021): 1985–97. http://dx.doi.org/10.3233/jcm215504.

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In order to solve the problem that the existing living cell microscopy technology can not display the detailed information of cells, a high sensitivity digital holographic living cell microscopy technology is proposed in this paper. By measuring the phase distribution and refractive index distribution of living cells, the data of living cells are extracted and converted into digital hologram of living cells. Simulation and comparison of the commonly used two-dimensional living cell microscope methods. The experimental results show that the high-sensitivity digital holographic microscopic detection method can obtain the detailed information of living cells, which proves the effectiveness of this study.

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Wait, Eric C., Michael A. Reiche und Teng-Leong Chew. „Hypothesis-driven quantitative fluorescence microscopy – the importance of reverse-thinking in experimental design“. Journal of Cell Science 133, Nr. 21 (01.11.2020): jcs250027. http://dx.doi.org/10.1242/jcs.250027.

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ABSTRACTOne of the challenges in modern fluorescence microscopy is to reconcile the conventional utilization of microscopes as exploratory instruments with their emerging and rapidly expanding role as a quantitative tools. The contribution of microscopy to observational biology will remain enormous owing to the improvements in acquisition speed, imaging depth, resolution and biocompatibility of modern imaging instruments. However, the use of fluorescence microscopy to facilitate the quantitative measurements necessary to challenge hypotheses is a relatively recent concept, made possible by advanced optics, functional imaging probes and rapidly increasing computational power. We argue here that to fully leverage the rapidly evolving application of microscopes in hypothesis-driven biology, we not only need to ensure that images are acquired quantitatively but must also re-evaluate how microscopy-based experiments are designed. In this Opinion, we present a reverse logic that guides the design of quantitative fluorescence microscopy experiments. This unique approach starts from identifying the results that would quantitatively inform the hypothesis and map the process backward to microscope selection. This ensures that the quantitative aspects of testing the hypothesis remain the central focus of the entire experimental design.

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Shotton, D. M. „Video-enhanced light microscopy and its applications in cell biology“. Journal of Cell Science 89, Nr. 2 (01.02.1988): 129–50. http://dx.doi.org/10.1242/jcs.89.2.129.

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The combination of novel optical microscopic techniques with advanced video and digital image-processing technology now permits dramatic improvements in the quality of light-microscope images. Such video-enhanced light microscopy has lead to a renaissance in the applications of the light microscope for the study of living cells in two important areas: the intensification of faint fluorescence images, permitting observation of fluorescently labelled cells under conditions of very low illuminating intensity; and the enhancement of extremely low contrast images generated by minute cellular structures, so that these may be clearly seen and their normal intracellular movements recorded. Application of both these aspects of video-enhanced light microscopy have recently led to major discoveries concerning the functioning of the living cell. In this review I discuss the equipment, procedures and image-processing principles employed in these applications, and describe and illustrate some of the spectacular results that have recently been obtained.

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Yang, Thomas Zhirui, und Yumin Wu. „Seeing cells without a lens: Compact 3D digital lensless holographic microscopy for wide-field imaging“. Theoretical and Natural Science 12, Nr. 1 (17.11.2023): 61–72. http://dx.doi.org/10.54254/2753-8818/12/20230434.

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Optical microscopy is an essential tool for biomedical discoveries and cell diagnosis at micro- to nano-scales. However, conventional microscopes rely on lenses to record 2-D images of samples, which limits in-depth inspection of large volumes of cells. This research project implements a novel 3-D lensless microscopic imaging system that achieves a wide field of view, high resolution, and an extremely compact, cost-effective design: the Digital Lensless Holographic Microscope (DLHM).A lensless holographic microscope is built with only a light source, a sample, and an imaging chip (with other non-essential supporting structures). The entire setup costs $500 to $600. A series of MATLAB-based algorithms were designed to reconstruct phase information of samples simultaneously from the recorded hologram with built-in high-resolution and phase unwrapping functions. This produces 3-D images of cell samples. The 3-D cell reconstruction of biological samples maintained a comparable resolution with conventional optical microscopes while covering a field of view of 36.2 mm2, which is 20-30 times larger. While most microscopes are extremely time-consuming and require professional expertise, the lensless holographic microscope is portable, low-cost, high-stability, and extremely simple. This makes it accessible for point-of-care testing (POCT) to a broader coverage, including developing regions with limited medical facilities.

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Kosaka, Yudai, und Tetsuhiko Ohba. „3P174 Study on membrane microfluidity of living cells using Muller Matrix microscopy(12. Cell biology,Poster)“. Seibutsu Butsuri 53, supplement1-2 (2013): S240. http://dx.doi.org/10.2142/biophys.53.s240_5.

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Schneckenburger, Herbert, und Christoph Cremer. „Axial Tomography in Live Cell Microscopy“. Biophysica 4, Nr. 2 (29.03.2024): 142–57. http://dx.doi.org/10.3390/biophysica4020010.

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For many biomedical applications, laser-assisted methods are essential to enhance the three-dimensional (3D) resolution of a light microscope. In this report, we review possibilities to improve the 3D imaging potential by axial tomography. This method allows us to rotate the object in a microscope into the best perspective required for imaging. Furthermore, images recorded under variable angles can be combined to one image with isotropic resolution. After a brief review of the technical state of the art, we show some biomedical applications, and discuss future perspectives for Deep View Microscopy and Molecular Imaging.

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Sagvolden, G., I. Giaever, E. O. Pettersen und J. Feder. „Cell adhesion force microscopy“. Proceedings of the National Academy of Sciences 96, Nr. 2 (19.01.1999): 471–76. http://dx.doi.org/10.1073/pnas.96.2.471.

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Konishi, Hiromi, Akira Ishikawa, Ying-Bing Jiang, Peter Buseck und Huifang Xu. „Sealed Environmental Cell Microscopy“. Microscopy and Microanalysis 9, S02 (15.07.2003): 902–3. http://dx.doi.org/10.1017/s1431927603444516.

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Hillebrand, Merle, Sophie E. Verrier, Andreas Ohlenbusch, Annika Schäfer, Hans-Dieter Söling, Fred S. Wouters und Jutta Gärtner. „Live Cell FRET Microscopy“. Journal of Biological Chemistry 282, Nr. 37 (03.07.2007): 26997–7005. http://dx.doi.org/10.1074/jbc.m702122200.

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Volkov, I. A., N. V. Frigo, L. F. Znamenskaya und O. R. Katunina. „Application of Confocal Laser Scanning Microscopy in Biology and Medicine“. Vestnik dermatologii i venerologii 90, Nr. 1 (24.02.2014): 17–24. http://dx.doi.org/10.25208/0042-4609-2014-90-1-17-24.

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Fluorescence confocal laser scanning microscopy and reflectance confocal laser scanning microscopy are up-to-date highend study methods. Confocal microscopy is used in cell biology and medicine. By using confocal microscopy, it is possible to study bioplasts and localization of protein molecules and other compounds relative to cell or tissue structures, and to monitor dynamic cell processes. Confocal microscopes enable layer-by-layer scanning of test items to create demonstrable 3D models. As compared to usual fluorescent microscopes, confocal microscopes are characterized by a higher contrast ratio and image definition.

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Wright, S. J., J. S. Walker, H. Schatten, C. Simerly, J. J. McCarthy und G. Schatten. „Confocal fluorescence microscopy with the tandem scanning light microscope“. Journal of Cell Science 94, Nr. 4 (01.12.1989): 617–24. http://dx.doi.org/10.1242/jcs.94.4.617.

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Applications of the tandem scanning confocal microscope (TSM) to fluorescence microscopy and its ability to resolve fluorescent biological structures are described. The TSM, in conjunction with a cooled charge-coupled device (cooled CCD) and conventional epifluorescence light source and filter sets, provided high-resolution, confocal data, so that different fluorescent cellular components were distinguished in three dimensions within the same cell. One of the unique features of the TSM is the ability to image fluorochromes excited by ultraviolet light (e.g. Hoechst, DAPI) in addition to fluorescein and rhodamine. Since the illumination is dim, photobleaching is insignificant and prolonged viewing of living specimens is possible. Series of optical sections taken in the Z-axis with the TSM were reproduced as stereo images and three-dimensional reconstructions. These data show that the TSM is potentially a powerful tool in fluorescence microscopy for determining three-dimensional relationships of complex structures within cells labeled with multiple fluorochromes.

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Carmichael, Stephen W., und Jon Charlesworth. „Correlating Fluorescence Microscopy with Electron Microscopy“. Microscopy Today 12, Nr. 1 (Januar 2004): 3–7. http://dx.doi.org/10.1017/s1551929500051749.

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The use of fluorescent probes is becoming more and more common in cell biology. It would be useful if we were able to correlate a fluorescent structure with an electron microscopic image. The ability to definitively identify a fluorescent organelle would be very valuable. Recently, Ying Ren, Michael Kruhlak, and David Bazett-Jones devised a clever technique to correlate a structure visualized in the light microscope, even a fluorescing cell, with transmission electron microscopy (TEM).Two keys to the technique of Ren et al are the use of grids (as used in the TEM) with widely spaced grid bars and the use of Quetol as the embedding resin. The grids allow for cells to be identified between the grid bars, and in turn the bars are used to keep the cell of interest in register throughout the processing for TEM. Quetol resin was used for embedding because of its low auto fluorescence and sectioning properties. The resin also becomes soft and can be cut and easily peeled from glass coverslips when heated to 70°C.

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Prabhakar, Neeraj, Markus Peurla, Olga Shenderova und Jessica M. Rosenholm. „Fluorescent and Electron-Dense Green Color Emitting Nanodiamonds for Single-Cell Correlative Microscopy“. Molecules 25, Nr. 24 (13.12.2020): 5897. http://dx.doi.org/10.3390/molecules25245897.

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Correlative light and electron microscopy (CLEM) is revolutionizing how cell samples are studied. CLEM provides a combination of the molecular and ultrastructural information about a cell. For the execution of CLEM experiments, multimodal fiducial landmarks are applied to precisely overlay light and electron microscopy images. Currently applied fiducials such as quantum dots and organic dye-labeled nanoparticles can be irreversibly quenched by electron beam exposure during electron microscopy. Generally, the sample is therefore investigated with a light microscope first and later with an electron microscope. A versatile fiducial landmark should offer to switch back from electron microscopy to light microscopy while preserving its fluorescent properties. Here, we evaluated green fluorescent and electron dense nanodiamonds for the execution of CLEM experiments and precisely correlated light microscopy and electron microscopy images. We demonstrated that green color emitting fluorescent nanodiamonds withstand electron beam exposure, harsh chemical treatments, heavy metal straining, and, importantly, their fluorescent properties remained intact for light microscopy.

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Mund, Markus, und Jonas Ries. „How good are my data? Reference standards in superresolution microscopy“. Molecular Biology of the Cell 31, Nr. 19 (01.09.2020): 2093–96. http://dx.doi.org/10.1091/mbc.e19-04-0189.

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Superresolution microscopy is becoming increasingly widespread in biological labs. While it holds enormous potential for biological discovery, it is a complex imaging technique that requires thorough optimization of various experimental parameters to yield data of the highest quality. Unfortunately, it remains challenging even for seasoned users to judge from the acquired images alone whether their superresolution microscopy pipeline is performing at its optimum, or if the image quality could be improved. Here, we describe how superresolution microscopists can objectively characterize their imaging pipeline using suitable reference standards, which are stereotypic so that the same structure can be imaged everywhere, every time, on every microscope. Quantitative analysis of reference standard images helps characterizing the performance of one’s own microscopes over time, allows objective benchmarking of newly developed microscopy and labeling techniques, and finally increases comparability of superresolution microscopy data between labs.

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Sharmin, Nazlee, Ava Chow und Alice Dong. „A Comparison Between Virtual and Conventional Microscopes in Health Science Education“. Canadian Journal of Learning and Technology 49, Nr. 2 (28.11.2023): 1–20. http://dx.doi.org/10.21432/cjlt28270.

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Virtual microscopes are computer or web-based programs that enable users to visualize digital slides and mimic the experience of using a real light microscope. Traditional light microscopes have always been an essential teaching tool in health science education to observe and learn cell and tissue structures. However, studies comparing virtual and real light microscopes in education reported learners’ satisfaction with virtual microscopes regarding their usability, image quality, efficiency, and availability. Although the use of virtual or web-based microscopy is increasing, there is no equivalent decrease in the number of schools utilizing traditional microscopes. We conducted a scoping review to investigate the comparative impact of conventional and virtual microscopes on different aspects of learning. We report a relative effect of virtual and light microscopy on student performance, long-term knowledge retention, and satisfaction. Our results show that virtual microscopy is superior to traditional microscopes as a teaching tool in health science education. Further studies are needed on different learning components to guide the best use of virtual microscopy as a sole teaching tool for health care education.

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Engels, F. M. „Developments in application of light and scanning electron microscopy techniques for cell wall degradation studies“. Netherlands Journal of Agricultural Science 44, Nr. 4 (01.12.1996): 357–73. http://dx.doi.org/10.18174/njas.v44i4.542.

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The results of recent technological developments in light and scanning electron microscopy closely used for research on forage cell wall degradation in ruminants, are reviewed. The indigestibility of forages by rumen microorganisms used to be ascribed mainly to an overall presence of lignin in the plant material. However, early light microscopic observations without application of histochemical staining revealed that some leaf and stem tissues were degraded completely. The early use of lignin detecting dyes, such as acid phloroglucinol or safranin, in light microscopy made it possible to discriminate between lignified undegradable and unlignified degradable plant tissues. The introduction of the scanning electron microscope enabled a further discrimination between degradable and undegradable cell wall and cell wall layers in plant tissues. As a result of continuous improvement of the techniques used in microscopy, e.g. section to slide, mirror sectioning, microspectrophotometry and cryo-ultramilling, forage indigestibility can now be attributed to the specific deposition and location of cutin/suberin or lignin layers inside the plant cell wall. These structural layers form barriers hindering access of rumen microorganisms to degradable parts of the cell wall.

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Kobayashi, Kan. „Fine structure of dendritic cells in the epithelial cell layer of the ox tongue“. Proceedings, annual meeting, Electron Microscopy Society of America 48, Nr. 3 (12.08.1990): 528–29. http://dx.doi.org/10.1017/s0424820100160194.

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It is known that some kinds of dendritic cells are distributed in the epithelial cell layer of mucous membrane consisting of stratified squamous epithelium. In the process of exfoliation of the epithelial layer from the underlying connective tissue, dendritic cell bodies exposed on the ruptured surface of the epithelium were detected by scanning electron microscopy. These cells were also observed by light microscopy as well as by transmission electron microscopy.Dorsal mucous membrane of the ox tongue was fixed in Karnovsky's fixative or in 10% formalin. For scanning electron microscopy samples were immersed in 3N-HCl solution for 2-3 weeks at room temperature. The epithelial cell layer was removed from the underlying connective tissue layer1). They were postfixed in tannic acid and then 1% OsO4 for 1 hr. After dehydration in an ethanol series, the specimens were dried by passing through t-butylalcohol, coated with platinum-palladium and observed under an S-800 scanning electron microscope. For transmission electron microscopy, small pieces of the fixed tissue were post-fixed in 1% OsO4 for 1.5 hr and then embedded in Araldite-Epon.

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Jost, Anna Payne-Tobin, und Jennifer C. Waters. „Designing a rigorous microscopy experiment: Validating methods and avoiding bias“. Journal of Cell Biology 218, Nr. 5 (20.03.2019): 1452–66. http://dx.doi.org/10.1083/jcb.201812109.

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Images generated by a microscope are never a perfect representation of the biological specimen. Microscopes and specimen preparation methods are prone to error and can impart images with unintended attributes that might be misconstrued as belonging to the biological specimen. In addition, our brains are wired to quickly interpret what we see, and with an unconscious bias toward that which makes the most sense to us based on our current understanding. Unaddressed errors in microscopy images combined with the bias we bring to visual interpretation of images can lead to false conclusions and irreproducible imaging data. Here we review important aspects of designing a rigorous light microscopy experiment: validation of methods used to prepare samples and of imaging system performance, identification and correction of errors, and strategies for avoiding bias in the acquisition and analysis of images.

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Grant, K. W., N. J. Anderson, J. A. Hammarback, A. Sweatt, B. Dawson, P. Moore und W. G. Jerome. „Laser Capture Microscopy as an Aid to Ultrastructural Analysis“. Microscopy and Microanalysis 6, S2 (August 2000): 842–43. http://dx.doi.org/10.1017/s1431927600036709.

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Laser capture microdissection (LCM) is a technique that provides homogenous cell populations for molecular and light microscopic analysis. During viewing by a standard wide-field microscope, a specific cell is selected. Heat from a near-infrared laser melts an ethylene vinyl acetate (EVA) transparent film which bonds to the individual selected cell. Several thousand cells can be selected and captured using this method. A homogeneous subpopulation of cells may be collected, one at a time, by histologic characteristics and/or histochemical staining from frozen sections, deparaffinized tissue, cell cultures or a blood smear.Previously, this technique has primarily been used to capture cells for DNA or RNA analysis. This study was undertaken to investigate the possibility of capturing a subpopulation of cultured cells in order to study their ultrastructure with the transmission electron microscope (TEM). We report here that cultured cells can be processed, captured and embedded for electron microscopy, in such a manner as to maintain ultrastructure.

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Shanmugapriya, Soundararajan Vijayarathna und Sreenivasan Sasidharan. „Functional Validation of DownRegulated MicroRNAs in HeLa Cells Treated with Polyalthia longifolia Leaf Extract Using Different Microscopic Approaches: A Morphological Alteration-Based Validation“. Microscopy and Microanalysis 25, Nr. 05 (06.08.2019): 1263–72. http://dx.doi.org/10.1017/s1431927619014776.

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AbstractSeveral microscopy methods have been developed to assess the morphological changes in cells in the investigations of the mode of cell death in response to a stimulus. Our recent finding on the treatment of the IC50 concentration (26.67 μg/mL) of Polyalthia longifolia leaf extract indicated the induction of apoptotic cell death via the regulation of miRNA in HeLa cells. Hence, the current study was conducted to validate the function of these downregulated microRNAs in P. longifolia-treated HeLa cells using microscopic approaches. These include scanning electron microscope (SEM), transmission electron microscope (TEM), and acridine orange/propidium iodide (AO/PI)-based fluorescent microscopy techniques by observing the morphological alterations to cells after transfection with mimic miRNA. Interestingly, the morphological changes observed in this study demonstrated the apoptotic hallmarks, for instance, cell blebbing, cell shrinkage, cytoplasmic and nuclear condensation, vacuolization, cytoplasmic extrusion, and the formation of apoptotic bodies, which proved the role of dysregulated miRNAs in apoptotic HeLa cell death after treatment with the P. longifolia leaf extract. Conclusively, the current study proved the crucial role of downregulated miR-484 and miR-221-5p in the induction of apoptotic cell death in P. longifolia-treated HeLa cells using three approaches—SEM, TEM, and AO/PI-based fluorescent microscope.

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Martin, Sonya, Antonio Virgilio Failla, Udo Spöri, Christoph Cremer und Ana Pombo. „Measuring the Size of Biological Nanostructures with Spatially Modulated Illumination Microscopy“. Molecular Biology of the Cell 15, Nr. 5 (Mai 2004): 2449–55. http://dx.doi.org/10.1091/mbc.e04-01-0045.

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Spatially modulated illumination fluorescence microscopy can in theory measure the sizes of objects with a diameter ranging between 10 and 200 nm and has allowed accurate size measurement of subresolution fluorescent beads (∼40–100 nm). Biological structures in this size range have so far been measured by electron microscopy. Here, we have labeled sites containing the active, hyperphosphorylated form of RNA polymerase II in the nucleus of HeLa cells by using the antibody H5. The spatially modulated illumination-microscope was compared with confocal laser scanning and electron microscopes and found to be suitable for measuring the size of cellular nanostructures in a biological setting. The hyperphosphorylated form of polymerase II was found in structures with a diameter of ∼70 nm, well below the 200-nm resolution limit of standard fluorescence microscopes.

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Paddock, S. W. „Tandem scanning reflected-light microscopy of cell-substratum adhesions and stress fibres in Swiss 3T3 cells“. Journal of Cell Science 93, Nr. 1 (01.05.1989): 143–46. http://dx.doi.org/10.1242/jcs.93.1.143.

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This paper describes two applications of the tandem scanning reflected-light microscope (TSM) for the observation of the structure of individual cells growing in tissue culture. First, the TSM is used as an alternative to interference reflection microscopy (IRM) or total internal reflection aqueous fluorescence microscopy (TIRAF) to observe cell-substratum adhesions in unstained living cells growing on a glass coverslip. Second, the TSM is used to produce improved images of cellular structures in 3T3 cells stained with various protein dyes including Napthol Blue Black (NBB) and Coomassie Brilliant Blue (CBB). More specifically, close contacts and focal contacts are resolved in living 3T3 cells, and features of the nucleus, the cytoskeleton and extracellular matrix are resolved in both NBB- and CBB-stained cells. The focal contacts and associated stress fibres are clearly imaged in NBB-stained cells. The TSM is an improvement over conventional incident light microscopy because of the confocal image excludes information from out-of-focus regions of the cytoplasm, and, unlike the laser-based confocal microscope, the actual colour of the specimen is viewed directly with TSM in almost real-time.

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Xianjun Zhang. „Development and Application of Cryogenic Optical Microscopy in Photosynthesis“. Acta Physica Sinica 73, Nr. 21 (2024): 0. http://dx.doi.org/10.7498/aps.73.20241072.

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Efficient photosynthesis reaction thanks to the flexible energy regulation of two important pigment-protein complexes photosystem II (PSII) and photosystem I (PSI). Cryogenic spectral microscopy provides information about the spatial distribution and physiological functional states of photosynthetic components in photosynthetic organisms. Under low temperatures, the uphill energy transfer between pigments is efficiently suppressed so that the temperature-dependent PSI can be well analyzed. Therefore, a cryogenic spectral microscope allows us to discuss the physiological events surrounding PSII and PSI in the independent microscopic zones. This technique can be used to complement the insufficiency of cryogenic electron microscopy and atomic force microscopy in analyzing the photophysics and photochemistry of photosynthetic species. Historically, cryogenic optical microscopes originated from the desire for single-molecule spectroscopy detection. Development to date, the combination of optical microscopies with various spectroscopic techniques has extended the possibility of a multi-perspective investigation in photosynthesis research. In this paper, I review the important and recent progress in cryogenic spectral microscopy in the field of natural photosynthesis research from two aspects: single-molecule spectroscopy and single-cell spectroscopy. Meanwhile, I illustrate the advantages of this technique in clarifying the correlation between structure variability and function of pigment-protein complexes, and the physiological responses of photosynthetic organisms to variable environments.

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Weinhardt, Venera, Jian-Hua Chen, Axel Ekman, Gerry McDermott, Mark A. Le Gros und Carolyn Larabell. „Imaging cell morphology and physiology using X-rays“. Biochemical Society Transactions 47, Nr. 2 (05.04.2019): 489–508. http://dx.doi.org/10.1042/bst20180036.

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Abstract Morphometric measurements, such as quantifying cell shape, characterizing sub-cellular organization, and probing cell–cell interactions, are fundamental in cell biology and clinical medicine. Until quite recently, the main source of morphometric data on cells has been light- and electron-based microscope images. However, many technological advances have propelled X-ray microscopy into becoming another source of high-quality morphometric information. Here, we review the status of X-ray microscopy as a quantitative biological imaging modality. We also describe the combination of X-ray microscopy data with information from other modalities to generate polychromatic views of biological systems. For example, the amalgamation of molecular localization data, from fluorescence microscopy or spectromicroscopy, with structural information from X-ray tomography. This combination of data from the same specimen generates a more complete picture of the system than that can be obtained by a single microscopy method. Such multimodal combinations greatly enhance our understanding of biology by combining physiological and morphological data to create models that more accurately reflect the complexities of life.

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Schneckenburger, Herbert. „Lasers in Live Cell Microscopy“. International Journal of Molecular Sciences 23, Nr. 9 (30.04.2022): 5015. http://dx.doi.org/10.3390/ijms23095015.

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Due to their unique properties—coherent radiation, diffraction limited focusing, low spectral bandwidth and in many cases short light pulses—lasers play an increasing role in live cell microscopy. Lasers are indispensable tools in 3D microscopy, e.g., confocal, light sheet or total internal reflection microscopy, as well as in super-resolution microscopy using wide-field or confocal methods. Further techniques, e.g., spectral imaging or fluorescence lifetime imaging (FLIM) often depend on the well-defined spectral or temporal properties of lasers. Furthermore, laser microbeams are used increasingly for optical tweezers or micromanipulation of cells. Three exemplary laser applications in live cell biology are outlined. They include fluorescence diagnosis, in particular in combination with Förster Resonance Energy Transfer (FRET), photodynamic therapy as well as laser-assisted optoporation, and demonstrate the potential of lasers in cell biology and—more generally—in biomedicine.

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Liao, Hong-Gang, und Haimei Zheng. „Liquid Cell Transmission Electron Microscopy“. Annual Review of Physical Chemistry 67, Nr. 1 (27.05.2016): 719–47. http://dx.doi.org/10.1146/annurev-physchem-040215-112501.

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Inoué, Shinya. „Video microscopy in cell biology“. Proceedings, annual meeting, Electron Microscopy Society of America 45 (August 1987): 632. http://dx.doi.org/10.1017/s0424820100127591.

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The application of video improves the image quality and extends the applicability of the light microscope to an unprecedented degree. Rectified, Plan Apochromatic high N.A. objectives can be used at full condenser N.A. in polarized light, D.I.C., single sideband edge enhanced, brightfield, or fluorescence, etc., microscopy to yield exceptionally well-corrected images with high resolution and shallow depth of field. Contrast and image features barely visible through the ocular are clearly displayed after analog and digital video enhancement. Stationary image blemishes and random noise due to low light levels can both be removed, and either recorded- or current-dynamic images can be projected for pseudocolor, monochrome, and stereo display. Serial optical sections recorded on laser disk recorders can be played back at different through-focusing speeds, or as through-focal stereo pairs, to reveal extraordinarily fine cellular details. Several examples of such applications in cell biology will be demonstrated at the meetings.

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Arun Anand, Arun Anand, und Bahram Javidi Bahram Javidi. „Digital holographic microscopy for automated 3D cell identification: an overview (Invited Paper)“. Chinese Optics Letters 12, Nr. 6 (2014): 060012–60017. http://dx.doi.org/10.3788/col201412.060012.

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Kinosita, K., H. Itoh, S. Ishiwata, K. Hirano, T. Nishizaka und T. Hayakawa. „Dual-view microscopy with a single camera: real-time imaging of molecular orientations and calcium.“ Journal of Cell Biology 115, Nr. 1 (01.10.1991): 67–73. http://dx.doi.org/10.1083/jcb.115.1.67.

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A new microscope technique, termed "W" (double view video) microscopy, enables simultaneous observation of two different images of an object through a single video camera or by eye. The image pair may, for example, be transmission and fluorescence, fluorescence at different wavelengths, or mutually perpendicular components of polarized fluorescence. Any video microscope can be converted into a dual imager by simple insertion of a small optical device. The continuous appearance of the dual image assures the best time resolution in existing and future video microscopes. As an application, orientations of actin protomers in individual, moving actin filaments have been imaged at the video rate. Asymmetric calcium influxes into a cell exposed to an intense electric pulse have also been visualized.

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White, J. G., W. B. Amos und M. Fordham. „An evaluation of confocal versus conventional imaging of biological structures by fluorescence light microscopy.“ Journal of Cell Biology 105, Nr. 1 (01.07.1987): 41–48. http://dx.doi.org/10.1083/jcb.105.1.41.

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Scanning confocal microscopes offer improved rejection of out-of-focus noise and greater resolution than conventional imaging. In such a microscope, the imaging and condenser lenses are identical and confocal. These two lenses are replaced by a single lens when epi-illumination is used, making confocal imaging particularly applicable to incident light microscopy. We describe the results we have obtained with a confocal system in which scanning is performed by moving the light beam, rather than the stage. This system is considerably faster than the scanned stage microscope and is easy to use. We have found that confocal imaging gives greatly enhanced images of biological structures viewed with epifluorescence. The improvements are such that it is possible to optically section thick specimens with little degradation in the image quality of interior sections.

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Baldock, Sara J., Abdullah C. S. Talari, Rachael Smith, Karen L. Wright und Lorna Ashton. „Single‐cell Raman microscopy of microengineered cell scaffolds“. Journal of Raman Spectroscopy 50, Nr. 3 (04.12.2018): 371–79. http://dx.doi.org/10.1002/jrs.5525.

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33

Wells, William A. „Lipid microscopy“. Journal of Cell Biology 175, Nr. 2 (16.10.2006): 196a. http://dx.doi.org/10.1083/jcb.1752rr3.

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Stelzer, Ernst H. K. „Optical microscopy“. Trends in Cell Biology 3, Nr. 9 (September 1993): 319–20. http://dx.doi.org/10.1016/0962-8924(93)90016-t.

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35

Amos, W. B. „Light Microscopy“. Trends in Cell Biology 3, Nr. 1 (Januar 1993): 28. http://dx.doi.org/10.1016/0962-8924(93)90199-b.

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36

Martin, Rick, und Soojung Shin. „Photomicroscopy Made Easy by Converting Cell Phones into “CellCams”“. American Biology Teacher 78, Nr. 1 (01.01.2016): 71–75. http://dx.doi.org/10.1525/abt.2016.78.1.71.

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Photo and video microscopy expand the utility of microscopic observations in education, but attachments needed for this have been prohibitively expensive or too fragile to allow students individual access to these techniques. We describe a do-it-yourself method using inexpensive materials that allows students to build an adaptor that will allow them to turn their cell phones into “CellCams” that they can use to capture microscopic images and videos. Activities are presented that give students ways to learn about concepts of microscopy using their self-collected images.

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Brama, Elisabeth, Christopher J. Peddie, Gary Wilkes, Yan Gu, Lucy M. Collinson und Martin L. Jones. „ultraLM and miniLM: Locator tools for smart tracking of fluorescent cells in correlative light and electron microscopy“. Wellcome Open Research 1 (13.12.2016): 26. http://dx.doi.org/10.12688/wellcomeopenres.10299.1.

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In-resin fluorescence (IRF) protocols preserve fluorescent proteins in resin-embedded cells and tissues for correlative light and electron microscopy, aiding interpretation of macromolecular function within the complex cellular landscape. Dual-contrast IRF samples can be imaged in separate fluorescence and electron microscopes, or in dual-modality integrated microscopes for high resolution correlation of fluorophore to organelle. IRF samples also offer a unique opportunity to automate correlative imaging workflows. Here we present two new locator tools for finding and following fluorescent cells in IRF blocks, enabling future automation of correlative imaging. The ultraLM is a fluorescence microscope that integrates with an ultramicrotome, which enables ‘smart collection’ of ultrathin sections containing fluorescent cells or tissues for subsequent transmission electron microscopy or array tomography. The miniLM is a fluorescence microscope that integrates with serial block face scanning electron microscopes, which enables ‘smart tracking’ of fluorescent structures during automated serial electron image acquisition from large cell and tissue volumes.

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Staehelin, L. Andrew, und Dominick J. Paolillo. „A brief history of how microscopic studies led to the elucidation of the 3D architecture and macromolecular organization of higher plant thylakoids“. Photosynthesis Research 145, Nr. 3 (September 2020): 237–58. http://dx.doi.org/10.1007/s11120-020-00782-3.

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Abstract Microscopic studies of chloroplasts can be traced back to the year 1678 when Antonie van Leeuwenhoek reported to the Royal Society in London that he saw green globules in grass leaf cells with his single-lens microscope. Since then, microscopic studies have continued to contribute critical insights into the complex architecture of chloroplast membranes and how their structure relates to function. This review is organized into three chronological sections: During the classic light microscope period (1678–1940), the development of improved microscopes led to the identification of green grana, a colorless stroma, and a membrane envelope. More recent (1990–2020) chloroplast dynamic studies have benefited from laser confocal and 3D-structured illumination microscopy. The development of the transmission electron microscope (1940–2000) and thin sectioning techniques demonstrated that grana consist of stacks of closely appressed grana thylakoids interconnected by non-appressed stroma thylakoids. When the stroma thylakoids were shown to spiral around the grana stacks as multiple right-handed helices, it was confirmed that the membranes of a chloroplast are all interconnected. Freeze-fracture and freeze-etch methods verified the helical nature of the stroma thylakoids, while also providing precise information on how the electron transport chain and ATP synthase complexes are non-randomly distributed between grana and stroma membrane regions. The last section (2000–2020) focuses on the most recent discoveries made possible by atomic force microscopy of hydrated membranes, and electron tomography and cryo-electron tomography of cryofixed thylakoids. These investigations have provided novel insights into thylakoid architecture and plastoglobules (summarized in a new thylakoid model), while also producing molecular-scale views of grana and stroma thylakoids in which individual functional complexes can be identified.

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Ma, Jianfeng, Zhe Ji, Xia Zhou, Zhiheng Zhang und Feng Xu. „Transmission Electron Microscopy, Fluorescence Microscopy, and Confocal Raman Microscopic Analysis of Ultrastructural and Compositional Heterogeneity of Cornus alba L. Wood Cell Wall“. Microscopy and Microanalysis 19, Nr. 1 (Februar 2013): 243–53. http://dx.doi.org/10.1017/s1431927612013906.

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AbstractTransmission electron microscopy (TEM), fluorescence microscopy, and confocal Raman microscopy can be used to characterize ultrastructural and compositional heterogeneity of plant cell walls. In this study, TEM observations revealed the ultrastructural characterization of Cornus alba L. fiber, vessel, axial parenchyma, ray parenchyma, and pit membrane between cells, notably with the ray parenchyma consisting of two well-defined layers. Fluorescence microscopy evidenced that cell corner middle lamella was more lignified than adjacent compound middle lamella and secondary wall with variation in lignification level from cell to cell. In situ Raman images showed that the inhomogeneity in cell wall components (cellulose and lignin) among different cells and within morphologically distinct cell wall layers. As the significant precursors of lignin biosynthesis, the pattern of coniferyl alcohol and aldehyde (joint abbreviation Lignin-CAA for both structures) distribution in fiber cell wall was also identified by Raman images, with higher concentration occurring in the fiber secondary wall where there was the highest cellulose concentration. Moreover, noteworthy was the observation that higher concentration of lignin and very minor amounts of cellulose were visualized in the pit membrane areas. These complementary microanalytical methods provide more accurate and complete information with regard to ultrastructural and compositional characterization of plant cell walls.

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Smith, R. W. „Non-Imaging Microscopies: Flow Cytometry as a Correlative Analytical Tool in the Quantification of Cell Structure, Autofluorescence, Fluorescent Probes and Cell Populations.“ Microscopy and Microanalysis 5, S2 (August 1999): 490–91. http://dx.doi.org/10.1017/s1431927600015774.

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Non-imaging microscopy has developed somewhat independently of both traditional light microscopy and laser confocal microscopy. Flow cytometry is the chief commercial and research technology among these microscopies, and, like other nonimaging detection systems, developed around the theme of automation in clinical laboratory medicine. It is an important correlative or parallel microscopy to several image forming microscopical methods. Cell sorting is an important option as well.The basic structure of the flow cytometer certainly parallels light, laser and electron microscopes. The flow cytometer has a light source, a set of adjustable optics to focus the beam on the specimen, objective optics to collect the light and direct it to appropriate sensors, and the sensors themselves. A real image is not formed because the sensors are not in an even plane with the projection, such as provided by the retina in light microscopy or an image plane or film plate in electron microscopy, and the objective optics may not focus in the image plane.While early flow cytometers were developed primarily for the automatic counting of cells and particles, modern instruments offer particular advantages for the analysis of fluorescence, fluorescent chemicals and probes and cellular auto fluorescence.

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Jiang, Nan, Hyeon-Jin Kim, Tyler J. Chozinski, Jorge E. Azpurua, Benjamin A. Eaton, Joshua C. Vaughan und Jay Z. Parrish. „Superresolution imaging of Drosophila tissues using expansion microscopy“. Molecular Biology of the Cell 29, Nr. 12 (15.06.2018): 1413–21. http://dx.doi.org/10.1091/mbc.e17-10-0583.

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The limited resolving power of conventional diffraction-limited microscopy hinders analysis of small, densely packed structural elements in cells. Expansion microscopy (ExM) provides an elegant solution to this problem, allowing for increased resolution with standard microscopes via physical expansion of the specimen in a swellable polymer hydrogel. Here, we apply, validate, and optimize ExM protocols that enable the study of Drosophila embryos, larval brains, and larval and adult body walls. We achieve a lateral resolution of ∼70 nm in Drosophila tissues using a standard confocal microscope, and we use ExM to analyze fine intracellular structures and intercellular interactions. First, we find that ExM reveals features of presynaptic active zone (AZ) structure that are observable with other superresolution imaging techniques but not with standard confocal microscopy. We further show that synapses known to exhibit age-dependent changes in activity also exhibit age-dependent changes in AZ structure. Finally, we use the significantly improved axial resolution of ExM to show that dendrites of somatosensory neurons are inserted into epithelial cells at a higher frequency than previously reported in confocal microscopy studies. Altogether, our study provides a foundation for the application of ExM to Drosophila tissues and underscores the importance of tissue-specific optimization of ExM procedures.

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O'Connell, Christopher B. „Live Cell Super-Resolution Imaging with N-SIM“. Microscopy Today 20, Nr. 4 (Juli 2012): 18–21. http://dx.doi.org/10.1017/s1551929512000375.

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The ability to visualize the distributions of specific proteins with a light microscope and fluorescent probes is largely responsible for our current understanding of cellular structure. A major limitation of this approach arises from the blurring effects of diffraction, which decreases resolution and limits the ability to obtain information at the nanoscale. There has been a tremendous drive to develop optical and computational methods that improve the resolution of the light microscope, and structured illumination microscopy (SIM) is one solution. This method uses patterned illumination to double both lateral and axial resolution. Nikon's N-SIM is a commercial system that integrates the most desirable features of light microscopy, specific labeling of molecules, and live cell imaging, with structured illumination. This provides the ability to achieve super resolution suitable for a range of biological applications.

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Roukos, Vassilis, Gianluca Pegoraro, Ty C. Voss und Tom Misteli. „Cell cycle staging of individual cells by fluorescence microscopy“. Nature Protocols 10, Nr. 2 (29.01.2015): 334–48. http://dx.doi.org/10.1038/nprot.2015.016.

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BULAT, TANJA, OTILIJA KETA, LELA KORIĆANAC, JELENA ŽAKULA, IVAN PETROVIĆ, ALEKSANDRA RISTIĆ-FIRA und DANIJELA TODOROVIĆ. „Radiation dose determines the method for quantification of DNA double strand breaks“. Anais da Academia Brasileira de Ciências 88, Nr. 1 (04.03.2016): 127–36. http://dx.doi.org/10.1590/0001-3765201620140553.

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ABSTRACT Ionizing radiation induces DNA double strand breaks (DSBs) that trigger phosphorylation of the histone protein H2AX (γH2AX). Immunofluorescent staining visualizes formation of γH2AX foci, allowing their quantification. This method, as opposed to Western blot assay and Flow cytometry, provides more accurate analysis, by showing exact position and intensity of fluorescent signal in each single cell. In practice there are problems in quantification of γH2AX. This paper is based on two issues: the determination of which technique should be applied concerning the radiation dose, and how to analyze fluorescent microscopy images obtained by different microscopes. HTB140 melanoma cells were exposed to γ-rays, in the dose range from 1 to 16 Gy. Radiation effects on the DNA level were analyzed at different time intervals after irradiation by Western blot analysis and immunofluorescence microscopy. Immunochemically stained cells were visualized with two types of microscopes: AxioVision (Zeiss, Germany) microscope, comprising an ApoTome software, and AxioImagerA1 microscope (Zeiss, Germany). Obtained results show that the level of γH2AX is time and dose dependent. Immunofluorescence microscopy provided better detection of DSBs for lower irradiation doses, while Western blot analysis was more reliable for higher irradiation doses. AxioVision microscope containing ApoTome software was more suitable for the detection of γH2AX foci.

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Morone, Nobuhiro, Eiji Usukura, Akihiro Narita und Jiro Usukura. „Improved unroofing protocols for cryo-electron microscopy, atomic force microscopy and freeze-etching electron microscopy and the associated mechanisms“. Microscopy 69, Nr. 6 (23.05.2020): 350–59. http://dx.doi.org/10.1093/jmicro/dfaa028.

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Abstract Unroofing, which is the mechanical shearing of a cell to expose the cytoplasmic surface of the cell membrane, is a unique preparation method that allows membrane cytoskeletons to be observed by cryo-electron microscopy, atomic force microscopy, freeze-etching electron microscopy and other methods. Ultrasound and adhesion have been known to mechanically unroof cells. In this study, unroofing using these two means was denoted sonication unroofing and adhesion unroofing, respectively. We clarified the mechanisms by which cell membranes are removed in these unroofing procedures and established efficient protocols for each based on the mechanisms. In sonication unroofing, fine bubbles generated by sonication adhered electrostatically to apical cell surfaces and then removed the apical (dorsal) cell membrane with the assistance of buoyancy and water flow. The cytoplasmic surface of the ventral cell membrane remaining on the grids became observable by this method. In adhesion unroofing, grids charged positively by coating with Alcian blue were pressed onto the cells, thereby tightly adsorbing the dorsal cell membrane. Subsequently, a part of the cell membrane strongly adhered to the grids was peeled from the cells and transferred onto the grids when the grids were lifted. This method thus allowed the visualization of the cytoplasmic surface of the dorsal cell membrane. This paper describes robust, improved protocols for the two unroofing methods in detail. In addition, micro-unroofing (perforation) likely due to nanobubbles is introduced as a new method to make cells transparent to electron beams.

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Ogenko, Volodymyr. „ACHIEVEMENTS IN PHYSICAL CHEMISTRY IN THE FIELD OF MICROSCOPY AND VISUALIZATION OF NANOSYSTEMS“. Ukrainian Chemistry Journal 89, Nr. 8 (29.09.2023): 63–77. http://dx.doi.org/10.33609/2708-129x.89.08.2023.63-77.

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The review presents modern views and the history of the development of microscopic studies of nanosystems which heve been started 2014, after the Nobel Prize in Chemistry was awarded to Eric Betzig, William Mörner, and Stefan Gell "for the development of super-resolved fluorescence microscopy". Their work ushered in a new era of optical microscopy, enabling the precise examination of individual molecules and molecular clusters by using optical microscopes. By circumventing the diffraction limitations that had constrained traditional optical microscopes, scientists gained access to the nanoscale realm, investigating structures within the 1–100 nanometer range. Special attention is paid to the use of carbon quantum dots and plasmon resonance to enhance fluorescence when obtaining the effect of super-resolution images, which allow the use of optical microscopes in the estimation of the sizes of cluster and single molecules. This breakthrough in removing the diffraction limitation allowed scientists to use the working range of 1–100 nm and obtain 3D images of nanosystems and images of living cells. Particular attention is paid to the achievements and prospects of high-resolution fluorescent nanoscopy SRM, which is successfully developing and studying the nanoworld in the range of 1–100 nm at the level of scanning electron microscopy. In cell biology, nanomedicine, etc. are developing roadmaps for scientific breakthroughs in super-resolution visualization methods for "live" images. Prospects of Immuno-SERS microscopy and medicine of individual diagnosis are considered Key Findings: This article highlights the achievements and future prospects of super-resolution fluorescence microscopy SRM. High-resolution fluorescence microscopy has proven instrumental in advancing our understanding of the living world within the 1–100 nanometer range, which is akin to the capabilities of scanning electron microscopy. Within the domains of cell biology and nanomedicine, roadmaps for scientific breakthroughs are emerging, fueled by super-resolution imaging techniques, providing "live" insights into cellular processes. The horizons of Immuno-SERS Microscopy and Personalized Diagnostics Medicine are expanding, promising exciting prospects in the field of medical diagnostics.

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Smith, Julie, Andreas Frøslev Mathisen, Nadja Funch Richardt, Ann-Sophie Vander Plaetsen, Filip Van Nieuwerburgh, Henrik Stender und Thore Hillig. „Feasibility of single-cell analysis of model cancer and foetal cells in blood after isolation by cell picking“. Tumor Biology 41, Nr. 2 (Februar 2019): 101042831882336. http://dx.doi.org/10.1177/1010428318823361.

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The objective of the present feasibility study was to transfer single cell line cells to either microscopy slides for downstream immune characterization or to polymerase chain reaction tubes for downstream DNA quantitation. Tumour cell lines, SKBR3 and MCF7 and trophoblast cell line JEG-3 were spiked in healthy donor blood. The CytoTrack system was used to scan the spiked blood samples to identify target cells. Individual target cells were identified, picked by use of a CytoPicker and deposited to either a microscopic slide or a polymerase chain reaction tube (PCR). Single tumour cells on microscopic slides were further immunostained with human epidermal growth factor receptor 2 (Her2) and epithelial cell adhesion molecule (EpCAM). From the picked cells in polymerase chain reaction tubes, DNA was amplified, quantified and used for Short Tandem Repeat genotyping. Depositing rare cells to microscopy slides was laborious with only five cells per hour. In this study with a trained operator, the picked cells had an 80.5% recovery rate. Depositing single trophoblast cells in PCR tubes was a faster process with 10 cells in 5 min. Immunostaining of isolated cells by both Her2 and EpCAM was possible but showed varying staining intensity. Presence of trophoblasts and contaminating white blood cells in PCR tubes after cell picking was confirmed based on DNA yield and mixed Short Tandem Repeat profiles in five out of eight samples. Using the CytoPicker tool, single tumour and trophoblast cells were successfully isolated and moved from blood samples, allowing subsequent immunostaining or Short Tandem Repeat genotyping.

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Kidoaki, Satoru, Kouske Hamano und Thasaneeya Kuboki. „GS1-3 TRACTION FORCE MICROSCOPY OF MESENCHYMAL STEM CELLS IN MODE OF FRUSTRATED DIFFERENTIATION(GS1: Cell and Tissue Biomechanics I)“. Proceedings of the Asian Pacific Conference on Biomechanics : emerging science and technology in biomechanics 2015.8 (2015): 118. http://dx.doi.org/10.1299/jsmeapbio.2015.8.118.

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Benchimol, Marlene. „Trichomonads under Microscopy“. Microscopy and Microanalysis 10, Nr. 5 (Oktober 2004): 528–50. http://dx.doi.org/10.1017/s1431927604040905.

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Trichomonads are flagellate protists, and among themTrichomonas vaginalisandTritrichomonas foetusare the most studied because they are parasites of the urogenital tract of humans and cattle, respectively. Microscopy provides new insights into the cell biology and morphology of these parasites, and thus allows better understanding of the main aspects of their physiology. Here, we review the ultrastructure ofT. foetusandT. vaginalis, stressing the participation of the axostyle in the process of cell division and showing that the pseudocyst may be a new form in the trichomonad cell cycle and not simply a degenerative form. Other organelles, such as the Golgi and hydrogenosomes, are also reviewed. The virus present in trichomonads is discussed.

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Steinbach, Arden. „Neutron microscopy“. Cell Biophysics 7, Nr. 1 (März 1985): 1–29. http://dx.doi.org/10.1007/bf02788636.

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