[{"year":"2022","acknowledgement":"This work was funded by the Austrian Science Fund (FWF) grant P31445 to F.K.M.S and the National Institute of Allergy and Infectious Diseases under awards R01AI147890 to R.A.D. This research was also supported by the Scientific Service Units (SSUs) of IST Austria through resources provided by Scientific Computing (SciComp), the Life Science Facility (LSF), and the Electron Microscopy Facility (EMF). We thank Dustin Morado for providing the software SubTOM for data processing. We also thank William Wan for critical reading of the manuscript and valuable feedback.","_id":"11155","oa_version":"Published Version","type":"journal_article","month":"06","abstract":[{"text":"The potential of energy filtering and direct electron detection for cryo-electron microscopy (cryo-EM) has been well documented. Here, we assess the performance of recently introduced hardware for cryo-electron tomography (cryo-ET) and subtomogram averaging (STA), an increasingly popular structural determination method for complex 3D specimens. We acquired cryo-ET datasets of EIAV virus-like particles (VLPs) on two contemporary cryo-EM systems equipped with different energy filters and direct electron detectors (DED), specifically a Krios G4, equipped with a cold field emission gun (CFEG), Thermo Fisher Scientific Selectris X energy filter, and a Falcon 4 DED; and a Krios G3i, with a Schottky field emission gun (XFEG), a Gatan Bioquantum energy filter, and a K3 DED. We performed constrained cross-correlation-based STA on equally sized datasets acquired on the respective systems. The resulting EIAV CA hexamer reconstructions show that both systems perform comparably in the 4–6 Å resolution range based on Fourier-Shell correlation (FSC). In addition, by employing a recently introduced multiparticle refinement approach, we obtained a reconstruction of the EIAV CA hexamer at 2.9 Å. Our results demonstrate the potential of the new generation of energy filters and DEDs for STA, and the effects of using different processing pipelines on their STA outcomes.","lang":"eng"}],"date_updated":"2023-08-03T06:25:23Z","date_created":"2022-04-15T07:10:26Z","file_date_updated":"2022-08-02T11:07:58Z","volume":214,"status":"public","external_id":{"pmid":["35351542"],"isi":["000790733600001"]},"citation":{"ista":"Obr M, Hagen WJH, Dick RA, Yu L, Kotecha A, Schur FK. 2022. Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. Journal of Structural Biology. 214(2), 107852.","mla":"Obr, Martin, et al. “Exploring High-Resolution Cryo-ET and Subtomogram Averaging Capabilities of Contemporary DEDs.” <i>Journal of Structural Biology</i>, vol. 214, no. 2, 107852, Elsevier, 2022, doi:<a href=\"https://doi.org/10.1016/j.jsb.2022.107852\">10.1016/j.jsb.2022.107852</a>.","apa":"Obr, M., Hagen, W. J. H., Dick, R. A., Yu, L., Kotecha, A., &#38; Schur, F. K. (2022). Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. <i>Journal of Structural Biology</i>. Elsevier. <a href=\"https://doi.org/10.1016/j.jsb.2022.107852\">https://doi.org/10.1016/j.jsb.2022.107852</a>","ama":"Obr M, Hagen WJH, Dick RA, Yu L, Kotecha A, Schur FK. Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. <i>Journal of Structural Biology</i>. 2022;214(2). doi:<a href=\"https://doi.org/10.1016/j.jsb.2022.107852\">10.1016/j.jsb.2022.107852</a>","short":"M. Obr, W.J.H. Hagen, R.A. Dick, L. Yu, A. Kotecha, F.K. Schur, Journal of Structural Biology 214 (2022).","ieee":"M. Obr, W. J. H. Hagen, R. A. Dick, L. Yu, A. Kotecha, and F. K. Schur, “Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs,” <i>Journal of Structural Biology</i>, vol. 214, no. 2. Elsevier, 2022.","chicago":"Obr, Martin, Wim J.H. Hagen, Robert A. Dick, Lingbo Yu, Abhay Kotecha, and Florian KM Schur. “Exploring High-Resolution Cryo-ET and Subtomogram Averaging Capabilities of Contemporary DEDs.” <i>Journal of Structural Biology</i>. Elsevier, 2022. <a href=\"https://doi.org/10.1016/j.jsb.2022.107852\">https://doi.org/10.1016/j.jsb.2022.107852</a>."},"intvolume":"       214","has_accepted_license":"1","publication_status":"published","oa":1,"acknowledged_ssus":[{"_id":"LifeSc"},{"_id":"ScienComp"},{"_id":"EM-Fac"}],"ddc":["570"],"date_published":"2022-06-01T00:00:00Z","department":[{"_id":"FlSc"}],"pmid":1,"publisher":"Elsevier","user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","short":"CC BY (4.0)"},"article_type":"original","article_processing_charge":"Yes (via OA deal)","scopus_import":"1","publication":"Journal of Structural Biology","file":[{"checksum":"0b1eb53447aae8e95ae4c12d193b0b00","file_id":"11722","date_updated":"2022-08-02T11:07:58Z","access_level":"open_access","date_created":"2022-08-02T11:07:58Z","file_name":"2022_JourStructuralBiology_Obr.pdf","success":1,"creator":"dernst","file_size":7080863,"content_type":"application/pdf","relation":"main_file"}],"day":"01","author":[{"last_name":"Obr","first_name":"Martin","id":"4741CA5A-F248-11E8-B48F-1D18A9856A87","full_name":"Obr, Martin"},{"first_name":"Wim J.H.","last_name":"Hagen","full_name":"Hagen, Wim J.H."},{"full_name":"Dick, Robert A.","first_name":"Robert A.","last_name":"Dick"},{"full_name":"Yu, Lingbo","last_name":"Yu","first_name":"Lingbo"},{"first_name":"Abhay","last_name":"Kotecha","full_name":"Kotecha, Abhay"},{"full_name":"Schur, Florian KM","id":"48AD8942-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0003-4790-8078","last_name":"Schur","first_name":"Florian KM"}],"article_number":"107852","title":"Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs","project":[{"grant_number":"P31445","name":"Structural conservation and diversity in retroviral capsid","call_identifier":"FWF","_id":"26736D6A-B435-11E9-9278-68D0E5697425"}],"issue":"2","language":[{"iso":"eng"}],"keyword":["Structural Biology"],"isi":1,"publication_identifier":{"issn":["1047-8477"]},"quality_controlled":"1","doi":"10.1016/j.jsb.2022.107852"},{"volume":213,"file_date_updated":"2021-11-15T13:11:27Z","date_created":"2021-11-15T12:21:42Z","date_updated":"2023-11-21T08:36:02Z","abstract":[{"text":"A precise quantitative description of the ultrastructural characteristics underlying biological mechanisms is often key to their understanding. This is particularly true for dynamic extra- and intracellular filamentous assemblies, playing a role in cell motility, cell integrity, cytokinesis, tissue formation and maintenance. For example, genetic manipulation or modulation of actin regulatory proteins frequently manifests in changes of the morphology, dynamics, and ultrastructural architecture of actin filament-rich cell peripheral structures, such as lamellipodia or filopodia. However, the observed ultrastructural effects often remain subtle and require sufficiently large datasets for appropriate quantitative analysis. The acquisition of such large datasets has been enabled by recent advances in high-throughput cryo-electron tomography (cryo-ET) methods. This also necessitates the development of complementary approaches to maximize the extraction of relevant biological information. We have developed a computational toolbox for the semi-automatic quantification of segmented and vectorized filamentous networks from pre-processed cryo-electron tomograms, facilitating the analysis and cross-comparison of multiple experimental conditions. GUI-based components simplify the processing of data and allow users to obtain a large number of ultrastructural parameters describing filamentous assemblies. We demonstrate the feasibility of this workflow by analyzing cryo-ET data of untreated and chemically perturbed branched actin filament networks and that of parallel actin filament arrays. In principle, the computational toolbox presented here is applicable for data analysis comprising any type of filaments in regular (i.e. parallel) or random arrangement. We show that it can ease the identification of key differences between experimental groups and facilitate the in-depth analysis of ultrastructural data in a time-efficient manner.","lang":"eng"}],"oa_version":"Published Version","month":"11","type":"journal_article","_id":"10290","acknowledgement":"This research was supported by the Scientific Service Units (SSUs) of IST Austria through resources provided by Scientific Computing (SciComp), the Life Science Facility (LSF), the BioImaging Facility (BIF), and the Electron Microscopy Facility (EMF). We also thank Victor-Valentin Hodirnau for help with cryo-ET data acquisition. The authors acknowledge support from IST Austria and from the Austrian Science Fund (FWF): M02495 to G.D. and Austrian Science Fund (FWF): P33367 to F.K.M.S.","year":"2021","date_published":"2021-11-03T00:00:00Z","ddc":["572"],"acknowledged_ssus":[{"_id":"ScienComp"},{"_id":"LifeSc"},{"_id":"Bio"},{"_id":"EM-Fac"}],"oa":1,"publication_status":"published","has_accepted_license":"1","intvolume":"       213","related_material":{"record":[{"relation":"software","id":"14502","status":"public"}]},"citation":{"ista":"Dimchev GA, Amiri B, Fäßler F, Falcke M, Schur FK. 2021. Computational toolbox for ultrastructural quantitative analysis of filament networks in cryo-ET data. Journal of Structural Biology. 213(4), 107808.","mla":"Dimchev, Georgi A., et al. “Computational Toolbox for Ultrastructural Quantitative Analysis of Filament Networks in Cryo-ET Data.” <i>Journal of Structural Biology</i>, vol. 213, no. 4, 107808, Elsevier , 2021, doi:<a href=\"https://doi.org/10.1016/j.jsb.2021.107808\">10.1016/j.jsb.2021.107808</a>.","apa":"Dimchev, G. A., Amiri, B., Fäßler, F., Falcke, M., &#38; Schur, F. K. (2021). Computational toolbox for ultrastructural quantitative analysis of filament networks in cryo-ET data. <i>Journal of Structural Biology</i>. Elsevier . <a href=\"https://doi.org/10.1016/j.jsb.2021.107808\">https://doi.org/10.1016/j.jsb.2021.107808</a>","ama":"Dimchev GA, Amiri B, Fäßler F, Falcke M, Schur FK. Computational toolbox for ultrastructural quantitative analysis of filament networks in cryo-ET data. <i>Journal of Structural Biology</i>. 2021;213(4). doi:<a href=\"https://doi.org/10.1016/j.jsb.2021.107808\">10.1016/j.jsb.2021.107808</a>","short":"G.A. Dimchev, B. Amiri, F. Fäßler, M. Falcke, F.K. Schur, Journal of Structural Biology 213 (2021).","ieee":"G. A. Dimchev, B. Amiri, F. Fäßler, M. Falcke, and F. K. Schur, “Computational toolbox for ultrastructural quantitative analysis of filament networks in cryo-ET data,” <i>Journal of Structural Biology</i>, vol. 213, no. 4. Elsevier , 2021.","chicago":"Dimchev, Georgi A, Behnam Amiri, Florian Fäßler, Martin Falcke, and Florian KM Schur. “Computational Toolbox for Ultrastructural Quantitative Analysis of Filament Networks in Cryo-ET Data.” <i>Journal of Structural Biology</i>. Elsevier , 2021. <a href=\"https://doi.org/10.1016/j.jsb.2021.107808\">https://doi.org/10.1016/j.jsb.2021.107808</a>."},"external_id":{"isi":["000720259500002"]},"status":"public","title":"Computational toolbox for ultrastructural quantitative analysis of filament networks in cryo-ET data","article_number":"107808","author":[{"last_name":"Dimchev","first_name":"Georgi A","orcid":"0000-0001-8370-6161","id":"38C393BE-F248-11E8-B48F-1D18A9856A87","full_name":"Dimchev, Georgi A"},{"full_name":"Amiri, Behnam","first_name":"Behnam","last_name":"Amiri"},{"id":"404F5528-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-7149-769X","full_name":"Fäßler, Florian","last_name":"Fäßler","first_name":"Florian"},{"full_name":"Falcke, Martin","first_name":"Martin","last_name":"Falcke"},{"first_name":"Florian KM","last_name":"Schur","full_name":"Schur, Florian KM","orcid":"0000-0003-4790-8078","id":"48AD8942-F248-11E8-B48F-1D18A9856A87"}],"day":"03","file":[{"access_level":"open_access","date_created":"2021-11-15T13:11:27Z","checksum":"6b209e4d44775d4e02b50f78982c15fa","file_id":"10291","date_updated":"2021-11-15T13:11:27Z","creator":"cchlebak","file_size":16818304,"relation":"main_file","content_type":"application/pdf","file_name":"2021_JournalStructBiol_Dimchev.pdf","success":1}],"publication":"Journal of Structural Biology","article_processing_charge":"Yes (via OA deal)","scopus_import":"1","article_type":"original","tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","short":"CC BY (4.0)"},"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","publisher":"Elsevier ","department":[{"_id":"FlSc"}],"doi":"10.1016/j.jsb.2021.107808","quality_controlled":"1","publication_identifier":{"issn":["1047-8477"]},"isi":1,"keyword":["Structural Biology"],"language":[{"iso":"eng"}],"issue":"4","project":[{"grant_number":"P33367","name":"Structure and isoform diversity of the Arp2/3 complex","_id":"9B954C5C-BA93-11EA-9121-9846C619BF3A"},{"grant_number":"M02495","name":"Protein structure and function in filopodia across scales","call_identifier":"FWF","_id":"2674F658-B435-11E9-9278-68D0E5697425"}]},{"citation":{"short":"F. Fäßler, B. Zens, R. Hauschild, F.K. Schur, Journal of Structural Biology 212 (2020).","chicago":"Fäßler, Florian, Bettina Zens, Robert Hauschild, and Florian KM Schur. “3D Printed Cell Culture Grid Holders for Improved Cellular Specimen Preparation in Cryo-Electron Microscopy.” <i>Journal of Structural Biology</i>. Elsevier, 2020. <a href=\"https://doi.org/10.1016/j.jsb.2020.107633\">https://doi.org/10.1016/j.jsb.2020.107633</a>.","ieee":"F. Fäßler, B. Zens, R. Hauschild, and F. K. Schur, “3D printed cell culture grid holders for improved cellular specimen preparation in cryo-electron microscopy,” <i>Journal of Structural Biology</i>, vol. 212, no. 3. Elsevier, 2020.","apa":"Fäßler, F., Zens, B., Hauschild, R., &#38; Schur, F. K. (2020). 3D printed cell culture grid holders for improved cellular specimen preparation in cryo-electron microscopy. <i>Journal of Structural Biology</i>. Elsevier. <a href=\"https://doi.org/10.1016/j.jsb.2020.107633\">https://doi.org/10.1016/j.jsb.2020.107633</a>","ista":"Fäßler F, Zens B, Hauschild R, Schur FK. 2020. 3D printed cell culture grid holders for improved cellular specimen preparation in cryo-electron microscopy. Journal of Structural Biology. 212(3), 107633.","mla":"Fäßler, Florian, et al. “3D Printed Cell Culture Grid Holders for Improved Cellular Specimen Preparation in Cryo-Electron Microscopy.” <i>Journal of Structural Biology</i>, vol. 212, no. 3, 107633, Elsevier, 2020, doi:<a href=\"https://doi.org/10.1016/j.jsb.2020.107633\">10.1016/j.jsb.2020.107633</a>.","ama":"Fäßler F, Zens B, Hauschild R, Schur FK. 3D printed cell culture grid holders for improved cellular specimen preparation in cryo-electron microscopy. <i>Journal of Structural Biology</i>. 2020;212(3). doi:<a href=\"https://doi.org/10.1016/j.jsb.2020.107633\">10.1016/j.jsb.2020.107633</a>"},"related_material":{"record":[{"status":"public","id":"14592","relation":"used_in_publication"},{"status":"public","id":"12491","relation":"dissertation_contains"}]},"intvolume":"       212","external_id":{"isi":["000600997800008"]},"status":"public","acknowledged_ssus":[{"_id":"ScienComp"},{"_id":"LifeSc"},{"_id":"Bio"},{"_id":"EM-Fac"}],"date_published":"2020-12-01T00:00:00Z","ddc":["570"],"has_accepted_license":"1","publication_status":"published","oa":1,"_id":"8586","year":"2020","acknowledgement":"This work was supported by the Austrian Science Fund (FWF, P33367) to FKMS. BZ acknowledges support by the Niederösterreich Fond. This research was also supported by the Scientific Service Units (SSU) of IST Austria through resources provided by Scientific Computing (SciComp), the Life Science Facility (LSF), the BioImaging Facility (BIF) and the Electron Microscopy Facility (EMF). We thank Georgi Dimchev (IST Austria) and Sonja Jacob (Vienna Biocenter Core Facilities) for testing our grid holders in different experimental setups and Daniel Gütl and the Kondrashov group (IST Austria) for granting us repeated access to their 3D printers. We also thank Jonna Alanko and the Sixt lab (IST Austria) for providing us HeLa cells, primary BL6 mouse tail fibroblasts, NIH 3T3 fibroblasts and human telomerase immortalised foreskin fibroblasts for our experiments. We are thankful to Ori Avinoam and William Wan for helpful comments on the manuscript and also thank Dorotea Fracchiolla (Art&Science) for illustrating the graphical abstract.","file_date_updated":"2020-12-10T14:01:10Z","date_created":"2020-09-29T13:24:06Z","volume":212,"month":"12","oa_version":"Published Version","type":"journal_article","abstract":[{"lang":"eng","text":"Cryo-electron microscopy (cryo-EM) of cellular specimens provides insights into biological processes and structures within a native context. However, a major challenge still lies in the efficient and reproducible preparation of adherent cells for subsequent cryo-EM analysis. This is due to the sensitivity of many cellular specimens to the varying seeding and culturing conditions required for EM experiments, the often limited amount of cellular material and also the fragility of EM grids and their substrate. Here, we present low-cost and reusable 3D printed grid holders, designed to improve specimen preparation when culturing challenging cellular samples directly on grids. The described grid holders increase cell culture reproducibility and throughput, and reduce the resources required for cell culturing. We show that grid holders can be integrated into various cryo-EM workflows, including micro-patterning approaches to control cell seeding on grids, and for generating samples for cryo-focused ion beam milling and cryo-electron tomography experiments. Their adaptable design allows for the generation of specialized grid holders customized to a large variety of applications."}],"date_updated":"2024-03-25T23:30:04Z","issue":"3","language":[{"iso":"eng"}],"keyword":["electron microscopy","cryo-EM","EM sample preparation","3D printing","cell culture"],"isi":1,"project":[{"grant_number":"P33367","name":"Structure and isoform diversity of the Arp2/3 complex","_id":"9B954C5C-BA93-11EA-9121-9846C619BF3A"},{"name":"NÖ-Fonds Preis für die Jungforscherin des Jahres am IST Austria","_id":"059B463C-7A3F-11EA-A408-12923DDC885E"}],"quality_controlled":"1","doi":"10.1016/j.jsb.2020.107633","publication_identifier":{"issn":["1047-8477"]},"tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","short":"CC BY (4.0)"},"article_type":"original","article_processing_charge":"Yes (via OA deal)","scopus_import":"1","publication":"Journal of Structural Biology","department":[{"_id":"FlSc"}],"user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","publisher":"Elsevier","article_number":"107633","title":"3D printed cell culture grid holders for improved cellular specimen preparation in cryo-electron microscopy","file":[{"content_type":"application/pdf","relation":"main_file","file_size":7076870,"creator":"dernst","success":1,"file_name":"2020_JourStrucBiology_Faessler.pdf","date_created":"2020-12-10T14:01:10Z","access_level":"open_access","date_updated":"2020-12-10T14:01:10Z","file_id":"8937","checksum":"c48cbf594e84fc2f91966ffaafc0918c"}],"day":"01","author":[{"last_name":"Fäßler","first_name":"Florian","orcid":"0000-0001-7149-769X","id":"404F5528-F248-11E8-B48F-1D18A9856A87","full_name":"Fäßler, Florian"},{"first_name":"Bettina","last_name":"Zens","full_name":"Zens, Bettina","id":"45FD126C-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Robert","last_name":"Hauschild","orcid":"0000-0001-9843-3522","id":"4E01D6B4-F248-11E8-B48F-1D18A9856A87","full_name":"Hauschild, Robert"},{"full_name":"Schur, Florian KM","orcid":"0000-0003-4790-8078","id":"48AD8942-F248-11E8-B48F-1D18A9856A87","last_name":"Schur","first_name":"Florian KM"}]},{"date_published":"2019-04-01T00:00:00Z","publication_status":"published","citation":{"ieee":"C. Bougault, I. Ayala, W. Vollmer, J.-P. Simorre, and P. Schanda, “Studying intact bacterial peptidoglycan by proton-detected NMR spectroscopy at 100 kHz MAS frequency,” <i>Journal of Structural Biology</i>, vol. 206, no. 1. Elsevier, pp. 66–72, 2019.","chicago":"Bougault, Catherine, Isabel Ayala, Waldemar Vollmer, Jean-Pierre Simorre, and Paul Schanda. “Studying Intact Bacterial Peptidoglycan by Proton-Detected NMR Spectroscopy at 100 kHz MAS Frequency.” <i>Journal of Structural Biology</i>. Elsevier, 2019. <a href=\"https://doi.org/10.1016/j.jsb.2018.07.009\">https://doi.org/10.1016/j.jsb.2018.07.009</a>.","short":"C. Bougault, I. Ayala, W. Vollmer, J.-P. Simorre, P. Schanda, Journal of Structural Biology 206 (2019) 66–72.","ama":"Bougault C, Ayala I, Vollmer W, Simorre J-P, Schanda P. Studying intact bacterial peptidoglycan by proton-detected NMR spectroscopy at 100 kHz MAS frequency. <i>Journal of Structural Biology</i>. 2019;206(1):66-72. doi:<a href=\"https://doi.org/10.1016/j.jsb.2018.07.009\">10.1016/j.jsb.2018.07.009</a>","mla":"Bougault, Catherine, et al. “Studying Intact Bacterial Peptidoglycan by Proton-Detected NMR Spectroscopy at 100 kHz MAS Frequency.” <i>Journal of Structural Biology</i>, vol. 206, no. 1, Elsevier, 2019, pp. 66–72, doi:<a href=\"https://doi.org/10.1016/j.jsb.2018.07.009\">10.1016/j.jsb.2018.07.009</a>.","ista":"Bougault C, Ayala I, Vollmer W, Simorre J-P, Schanda P. 2019. Studying intact bacterial peptidoglycan by proton-detected NMR spectroscopy at 100 kHz MAS frequency. Journal of Structural Biology. 206(1), 66–72.","apa":"Bougault, C., Ayala, I., Vollmer, W., Simorre, J.-P., &#38; Schanda, P. (2019). Studying intact bacterial peptidoglycan by proton-detected NMR spectroscopy at 100 kHz MAS frequency. <i>Journal of Structural Biology</i>. Elsevier. <a href=\"https://doi.org/10.1016/j.jsb.2018.07.009\">https://doi.org/10.1016/j.jsb.2018.07.009</a>"},"intvolume":"       206","extern":"1","external_id":{"pmid":["30031884"]},"status":"public","date_created":"2020-09-17T10:29:10Z","volume":206,"month":"04","type":"journal_article","oa_version":"Submitted Version","abstract":[{"lang":"eng","text":"The bacterial cell wall is composed of the peptidoglycan (PG), a large polymer that maintains the integrity of the bacterial cell. Due to its multi-gigadalton size, heterogeneity, and dynamics, atomic-resolution studies are inherently complex. Solid-state NMR is an important technique to gain insight into its structure, dynamics and interactions. Here, we explore the possibilities to study the PG with ultra-fast (100 kHz) magic-angle spinning NMR. We demonstrate that highly resolved spectra can be obtained, and show strategies to obtain site-specific resonance assignments and distance information. We also explore the use of proton-proton correlation experiments, thus opening the way for NMR studies of intact cell walls without the need for isotope labeling."}],"date_updated":"2021-01-12T08:19:05Z","page":"66-72","_id":"8409","year":"2019","quality_controlled":"1","doi":"10.1016/j.jsb.2018.07.009","publication_identifier":{"issn":["1047-8477"]},"issue":"1","language":[{"iso":"eng"}],"keyword":["Structural Biology"],"title":"Studying intact bacterial peptidoglycan by proton-detected NMR spectroscopy at 100 kHz MAS frequency","day":"01","author":[{"last_name":"Bougault","first_name":"Catherine","full_name":"Bougault, Catherine"},{"full_name":"Ayala, Isabel","last_name":"Ayala","first_name":"Isabel"},{"first_name":"Waldemar","last_name":"Vollmer","full_name":"Vollmer, Waldemar"},{"full_name":"Simorre, Jean-Pierre","last_name":"Simorre","first_name":"Jean-Pierre"},{"full_name":"Schanda, Paul","id":"7B541462-FAF6-11E9-A490-E8DFE5697425","orcid":"0000-0002-9350-7606","first_name":"Paul","last_name":"Schanda"}],"article_type":"original","article_processing_charge":"No","publication":"Journal of Structural Biology","pmid":1,"publisher":"Elsevier","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87"},{"_id":"9655","year":"2014","file_date_updated":"2021-07-22T08:06:34Z","date_created":"2021-07-14T09:05:42Z","volume":186,"date_updated":"2021-07-22T08:26:32Z","abstract":[{"text":"Correlative microscopy incorporates the specificity of fluorescent protein labeling into high-resolution electron micrographs. Several approaches exist for correlative microscopy, most of which have used the green fluorescent protein (GFP) as the label for light microscopy. Here we use chemical tagging and synthetic fluorophores instead, in order to achieve protein-specific labeling, and to perform multicolor imaging. We show that synthetic fluorophores preserve their post-embedding fluorescence in the presence of uranyl acetate. Post-embedding fluorescence is of such quality that the specimen can be prepared with identical protocols for scanning electron microscopy (SEM) and transmission electron microscopy (TEM); this is particularly valuable when singular or otherwise difficult samples are examined. We show that synthetic fluorophores give bright, well-resolved signals in super-resolution light microscopy, enabling us to superimpose light microscopic images with a precision of up to 25 nm in the x–y plane on electron micrographs. To exemplify the preservation quality of our new method we visualize the molecular arrangement of cadherins in adherens junctions of mouse epithelial cells.","lang":"eng"}],"oa_version":"Published Version","type":"journal_article","month":"05","page":"205-213","citation":{"short":"M. Perkovic, M. Kunz, U. Endesfelder, S. Bunse, C. Wigge, Z. Yu, V.-V. Hodirnau, M.P. Scheffer, A. Seybert, S. Malkusch, E.M. Schuman, M. Heilemann, A.S. Frangakis, Journal of Structural Biology 186 (2014) 205–213.","ieee":"M. Perkovic <i>et al.</i>, “Correlative light- and electron microscopy with chemical tags,” <i>Journal of Structural Biology</i>, vol. 186, no. 2. Elsevier, pp. 205–213, 2014.","chicago":"Perkovic, Mario, Michael Kunz, Ulrike Endesfelder, Stefanie Bunse, Christoph Wigge, Zhou Yu, Victor-Valentin Hodirnau, et al. “Correlative Light- and Electron Microscopy with Chemical Tags.” <i>Journal of Structural Biology</i>. Elsevier, 2014. <a href=\"https://doi.org/10.1016/j.jsb.2014.03.018\">https://doi.org/10.1016/j.jsb.2014.03.018</a>.","mla":"Perkovic, Mario, et al. “Correlative Light- and Electron Microscopy with Chemical Tags.” <i>Journal of Structural Biology</i>, vol. 186, no. 2, Elsevier, 2014, pp. 205–13, doi:<a href=\"https://doi.org/10.1016/j.jsb.2014.03.018\">10.1016/j.jsb.2014.03.018</a>.","ista":"Perkovic M, Kunz M, Endesfelder U, Bunse S, Wigge C, Yu Z, Hodirnau V-V, Scheffer MP, Seybert A, Malkusch S, Schuman EM, Heilemann M, Frangakis AS. 2014. Correlative light- and electron microscopy with chemical tags. Journal of Structural Biology. 186(2), 205–213.","apa":"Perkovic, M., Kunz, M., Endesfelder, U., Bunse, S., Wigge, C., Yu, Z., … Frangakis, A. S. (2014). Correlative light- and electron microscopy with chemical tags. <i>Journal of Structural Biology</i>. Elsevier. <a href=\"https://doi.org/10.1016/j.jsb.2014.03.018\">https://doi.org/10.1016/j.jsb.2014.03.018</a>","ama":"Perkovic M, Kunz M, Endesfelder U, et al. Correlative light- and electron microscopy with chemical tags. <i>Journal of Structural Biology</i>. 2014;186(2):205-213. doi:<a href=\"https://doi.org/10.1016/j.jsb.2014.03.018\">10.1016/j.jsb.2014.03.018</a>"},"extern":"1","intvolume":"       186","external_id":{"pmid":["24698954"]},"status":"public","ddc":["570"],"date_published":"2014-05-01T00:00:00Z","has_accepted_license":"1","publication_status":"published","oa":1,"article_processing_charge":"No","scopus_import":"1","article_type":"original","tmp":{"name":"Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported (CC BY-NC-ND 3.0)","short":"CC BY-NC-ND (3.0)","legal_code_url":"https://creativecommons.org/licenses/by-nc-nd/3.0/legalcode","image":"/images/cc_by_nc_nd.png"},"publication":"Journal of Structural Biology","pmid":1,"publisher":"Elsevier","user_id":"6785fbc1-c503-11eb-8a32-93094b40e1cf","title":"Correlative light- and electron microscopy with chemical tags","file":[{"file_size":3454628,"relation":"main_file","content_type":"application/pdf","creator":"asandaue","file_name":"2014_JournalOfStructuralBiology_Perkovic.pdf","success":1,"date_created":"2021-07-22T08:06:34Z","access_level":"open_access","file_id":"9701","date_updated":"2021-07-22T08:06:34Z","checksum":"a322991b43cdc5935c99db88d285aa3a"}],"license":"https://creativecommons.org/licenses/by-nc-nd/3.0/","day":"01","author":[{"full_name":"Perkovic, Mario","last_name":"Perkovic","first_name":"Mario"},{"full_name":"Kunz, Michael","first_name":"Michael","last_name":"Kunz"},{"full_name":"Endesfelder, Ulrike","last_name":"Endesfelder","first_name":"Ulrike"},{"first_name":"Stefanie","last_name":"Bunse","full_name":"Bunse, Stefanie"},{"first_name":"Christoph","last_name":"Wigge","full_name":"Wigge, Christoph"},{"last_name":"Yu","first_name":"Zhou","full_name":"Yu, Zhou"},{"id":"3661B498-F248-11E8-B48F-1D18A9856A87","full_name":"Hodirnau, Victor-Valentin","last_name":"Hodirnau","first_name":"Victor-Valentin"},{"first_name":"Margot P.","last_name":"Scheffer","full_name":"Scheffer, Margot P."},{"full_name":"Seybert, Anja","last_name":"Seybert","first_name":"Anja"},{"first_name":"Sebastian","last_name":"Malkusch","full_name":"Malkusch, Sebastian"},{"last_name":"Schuman","first_name":"Erin M.","full_name":"Schuman, Erin M."},{"first_name":"Mike","last_name":"Heilemann","full_name":"Heilemann, Mike"},{"first_name":"Achilleas S.","last_name":"Frangakis","full_name":"Frangakis, Achilleas S."}],"language":[{"iso":"eng"}],"issue":"2","quality_controlled":"1","doi":"10.1016/j.jsb.2014.03.018","publication_identifier":{"issn":["1047-8477"]}}]