Готові списки джерел за темами / Cryo-CLEM

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Добірка наукової літератури з теми "Cryo-CLEM"

Автор: Grafiati

Опубліковано: 14 вересня 2024

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Статті в журналах з теми "Cryo-CLEM"

Metskas, Lauren Ann, and John A. G. Briggs. "Fluorescence-Based Detection of Membrane Fusion State on a Cryo-EM Grid using Correlated Cryo-Fluorescence and Cryo-Electron Microscopy." Microscopy and Microanalysis 25, no. 4 (May 14, 2019): 942–49. http://dx.doi.org/10.1017/s1431927619000606.

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AbstractCorrelated light and electron microscopy (CLEM) has become a popular technique for combining the protein-specific labeling of fluorescence with electron microscopy, both at room and cryogenic temperatures. Fluorescence applications at cryo-temperatures have typically been limited to localization of tagged protein oligomers due to known issues of extended triplet state duration, spectral shifts, and reduced photon capture through cryo-CLEM objectives. Here, we consider fluorophore characteristics and behaviors that could enable more extended applications. We describe how dialkylcarbocanine DiD, and its autoquenching by resonant energy transfer (RET), can be used to distinguish the fusion state of a lipid bilayer at cryo-temperatures. By adapting an established fusion assay to work under cryo-CLEM conditions, we identified areas of fusion between influenza virus-like particles and fluorescently labeled lipid vesicles on a cryo-EM grid. This result demonstrates that cryo-CLEM can be used to localize functions in addition to tagged proteins, and that fluorescence autoquenching by RET can be incorporated successfully into cryo-CLEM approaches. In the case of membrane fusion applications, this method provides both an orthogonal confirmation of functional state independent of the morphological description from cryo-EM and a way to bridge room-temperature kinetic assays and the cryo-EM images.

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Moser, Felipe, Vojtěch Pražák, Valerie Mordhorst, Débora M. Andrade, Lindsay A. Baker, Christoph Hagen, Kay Grünewald, and Rainer Kaufmann. "Cryo-SOFI enabling low-dose super-resolution correlative light and electron cryo-microscopy." Proceedings of the National Academy of Sciences 116, no. 11 (February 26, 2019): 4804–9. http://dx.doi.org/10.1073/pnas.1810690116.

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Correlative light and electron cryo-microscopy (cryo-CLEM) combines information from the specific labeling of fluorescence cryo-microscopy (cryo-FM) with the high resolution in environmental context of electron cryo-microscopy (cryo-EM). Exploiting super-resolution methods for cryo-FM is advantageous, as it enables the identification of rare events within the environmental background of cryo-EM at a sensitivity and resolution beyond that of conventional methods. However, due to the need for relatively high laser intensities, current super-resolution cryo-CLEM methods require cryo-protectants or support films which can severely reduce image quality in cryo-EM and are not compatible with many samples, such as mammalian cells. Here, we introduce cryogenic super-resolution optical fluctuation imaging (cryo-SOFI), a low-dose super-resolution imaging scheme based on the SOFI principle. As cryo-SOFI does not require special sample preparation, it is fully compatible with conventional cryo-EM specimens, and importantly, it does not affect the quality of cryo-EM imaging. By applying cryo-SOFI to a variety of biological application examples, we demonstrate resolutions up to ∼135 nm, an improvement of up to three times compared with conventional cryo-FM, while maintaining the specimen in a vitrified state for subsequent cryo-EM. Cryo-SOFI presents a general solution to the problem of specimen devitrification in super-resolution cryo-CLEM. It does not require a complex optical setup and can easily be implemented in any existing cryo-FM system.

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Li, Shuoguo, Gang Ji, Xiaojun Huang, Lei Sun, Jianguo Zhang, Wei Xu, and Fei Sun. "A New Solution of Non-integrated Correlative Light and Electron Microscopy Based on High-vacuum Optical Platform." Microscopy and Microanalysis 22, S3 (July 2016): 248–49. http://dx.doi.org/10.1017/s1431927616002099.

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Abstract Correlative light and electron microscopy (CLEM) offers a means of guiding the search for the unique or rare events by fluorescence microscopy (FM) and allows electron microscopy (EM) to zoom in on them for subsequent EM examination in three-dimensions (3D) and with nanometer-scale resolution. FM visualizes the localization of specific antigens by using fluorescent tags or proteins in a large field-of-view to study their cellular function, whereas EM provides the high level of resolution for complex structures. And cryo CLEM combines the advantages of maintaining structural preservation in a near-native state throughout the entire imaging process and by avoiding potentially harmful pre-treatments, such as chemical fixation, dehydration and staining with heavy metals. Besides for frozen-hydrated biological samples, CLEM combines the advantages of a close-to-life preservation of biological materials by keeping them embedded in vitreous ice throughout the entire imaging process and the frozen-hydrated condition is very suitable to maintain fluorescent signals. In recent years, many new instruments and software which intended to optimize the workflow and to obtain better experimental results of CLEM have been presented or even commoditized. While, the specimen damage during transfer from FM to EM and the resolution of CLEM were still need to be improved. Here we set up a High-vacuum Optical Platform to develop CLEM imaging technology (HOPE), which was designed to realize high-vacuum optical ( fluorescent) imaging for cryo-sample on EM cryo-holder (e.g. Gatan 626). A non-integrated high-vacuum cryo-optical stage, which adapted to the EM cryo holder, was fixed on epi-fluorescence microscope (or super-resolution microscope) to obtain fluorescent images. And then the EM cryo holder would be transferred to EM for collection of EM data. This protocol was aimed to minimize the specimen damage during transfer from FM to EM and it was versatile to expend to different types of light microscopy or electron microscopy. Our HOPE had already passed correlative imaging test, and the results showed that it was convenient and effective.

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Hampton, Cheri M. "Practical Strategies for cryo-CLEM Experiments." Microscopy and Microanalysis 23, S1 (July 2017): 1400–1401. http://dx.doi.org/10.1017/s1431927617007668.

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Thomas, Connon I., Nicolai T. Urban, Ye Sun, Lesley A. Colgan, Xun Tu, Ryohei Yasuda, and Naomi Kamasawa. "Cryo-Confocal Imaging for CLEM Mapping in Brain Tissues." Microscopy Today 29, no. 5 (September 2021): 34–39. http://dx.doi.org/10.1017/s1551929521001073.

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Abstract:In correlative light and electron microscopy (CLEM) workflows, identifying the same sub-cellular features in tissue by both light (LM) and electron microscopy (EM) remains a challenge. Furthermore, use of cryo-fixation for EM is desirable to capture rapid biological phenomena. Here, we describe a workflow that incorporates cryo-confocal laser scanning microscopy into the CLEM process, mapping cells in brain slices to re-image them with serial section scanning electron microscopy (ssSEM) array tomography. The addition of Airyscan detection increased the signal-to-noise ratio (SNR), allowing individual spines in thick frozen tissue to be visualized at a sufficient spatial resolution, providing a new tool for a CLEM approach to capture biological dynamics.

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Kamp, Arnold, Martijn van Nugteren, Hildo Vader, Michael Schwertner, Duncan Stacey, Roman Koning, and Bram Koster. "Automated Cryo-plunging Robot to Prepare Samples for Single Particle Analysis (SPA), Cryo-EM, Cryo-ET, Cryo-fluorescence and Cryo-CLEM." Microscopy and Microanalysis 26, S2 (July 30, 2020): 2732–33. http://dx.doi.org/10.1017/s1431927620022606.

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Paraan, Reza, Victoria Hewitt, Yusuke Hirabayashi, Franck Polleux, Clint Potter, and Bridget Carragher. "Characterization of ER-mitochondria contact sites using cryo-CLEM." Microscopy and Microanalysis 27, S1 (July 30, 2021): 1712–13. http://dx.doi.org/10.1017/s1431927621006255.

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Schwertner, Michael, and Duncan Stacey. "Cryo-Correlative Light and Electron Microscopy (Cryo-CLEM): Specimen Workflow Paths and Recent Instrument Developments." Microscopy and Microanalysis 21, S3 (August 2015): 1565–66. http://dx.doi.org/10.1017/s1431927615008600.

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Sexton, Danielle L., Steffen Burgold, Andreas Schertel, and Elitza I. Tocheva. "Super-resolution confocal cryo-CLEM with cryo-FIB milling for in situ imaging of Deinococcus radiodurans." Current Research in Structural Biology 4 (2022): 1–9. http://dx.doi.org/10.1016/j.crstbi.2021.12.001.

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Shahmoradian, Sarah, Jim Monistrol, Hung Tri Tran, Valerie Perez, Jenny Jiou, Jaime Vaquer-Alicea, and Marc Diamond. "Abstract 2445 Elucidating Tau Fibril Formation using Correlative Cryo-CLEM in situ." Journal of Biological Chemistry 300, no. 3 (March 2024): 107098. http://dx.doi.org/10.1016/j.jbc.2024.107098.

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Більше джерел

Дисертації з теми "Cryo-CLEM"

Patra, Gurudatt. "Structure of mitotic chromosome and the role of condensin protein in the structural organization of chromosomes." Electronic Thesis or Diss., Strasbourg, 2024. http://www.theses.fr/2024STRAJ020.

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Au cours de la mitose, la chromatine interphasique subit une compaction massive en structures en forme de bâtonnets. Les condensines sont des complexes protéiques dont on sait qu'ils jouent un rôle majeur dans l'organisation des chromosomes mitotiques. Les eucaryotes possèdent deux complexes de condensines conservés, à savoir les condensines 1 et 2. Des études in vitro sur des modèles d'ADN nus montrent que les condensines ont une activité d'extrusion de boucles dans l'organisation des chromosomes. Cependant, il reste encore beaucoup à explorer en ce qui concerne l'étude de la fonction des condensines dans l'environnement encombré de la chromatine. Nous avons utilisé la technologie halo tag où le domaine SMC2 des condensines est marqué par fluorescence à l'aide d'un ligand halo TMR. Cette approche nous aide à localiser les régions riches en condensines dans les chromosomes mitotiques partiellement décondensés en utilisant la cryo-microscopie en lumière à l'intérieur des chromosomes vitrifiés pour les études de cryo-tomographie électronique. Nos tomographies montrent les complexes de condensine dans l'environnement chromatinien. Cela ouvre une fenêtre sur l'étude de l'activité de liaison à l'ADN de la condensine, l'oligomérisation ou le regroupement de la condensine et son interaction avec d'autres composants non histoniques des chromosomes mitotiques
During mitosis, the interphase chromatin undergoes a massive round of compaction into rod-shaped structures. Condensins are protein complexes that have been known to play a major role in mitotic chromosome organization. Eukaryotes have two conserved condensin complexes, namely condensin 1 and 2. In vitro studies on naked DNA templates show evidence for loop extrusion activity of condensins in chromosome organization. However, there is still a lot to explore regarding the study of condensin function inside the crowded chromatin environment. We have used halo tag technology where the SMC2 domain of condensins is tagged to fluorescently label using a halo TMR ligand. This approach helps us to locate condensin-rich regions in partially decondensed mitotic chromosomes using cryo-light microscopy inside the vitrified chromosomes for cryo-electron tomography studies. Our tomograms show condensin complexes inside the chromatin environment. This opens up a window into the study of DNA binding activity of condensin, the oligomerization or clustering of condensin and its interaction with other non-histone components of mitotic chromosomes

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Nocera, Giovanni Marco. "Microfluidic cryofixation for time-correlated live-imaging cryo-fluorescence microscopy and electron microscopy of Caenorhabditis elegans." Doctoral thesis, 2018. http://hdl.handle.net/11858/00-1735-0000-002E-E4EA-8.

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Частини книг з теми "Cryo-CLEM"

Zanetti-Domingues, Laura C., Michael Hirsch, Lin Wang, Tara A. Eastwood, Karen Baker, Daniel P. Mulvihill, Sheena Radford, Jim Horne, Paul White, and Benji Bateman. "Toward quantitative super-resolution methods for cryo-CLEM." In Methods in Cell Biology. Elsevier, 2024. http://dx.doi.org/10.1016/bs.mcb.2024.02.028.

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Тези доповідей конференцій з теми "Cryo-CLEM"

Smeets, Marit. "METEOR: an integrated top down cryo-CLEM imaging system." In Microscience Microscopy Congress 2021 incorporating EMAG 2021. Royal Microscopical Society, 2021. http://dx.doi.org/10.22443/rms.mmc2021.59.

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Schwertner, Michael. "Automated cryo-plunger for the preparation of vitrified cryo-samples for Single Particle Analysis (SPA), cryo-EM, cryo-ET and cryo-CLEM, based on a novel procedure replacing conventional blotting." In European Microscopy Congress 2020. Royal Microscopical Society, 2021. http://dx.doi.org/10.22443/rms.emc2020.1112.

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Schilling, Nicolas. "A cryo-preparation approach for immuno-fluorescence labelling of cell cultures and CLEM." In European Microscopy Congress 2020. Royal Microscopical Society, 2021. http://dx.doi.org/10.22443/rms.emc2020.867.

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