Article
Chemistry, Multidisciplinary
Michael Holtmannspoetter, Eike Wienbeuker, Timo Dellmann, Isabelle Watrinet, Ana J. Garcia-Saez, Kai Johnsson, Rainer Kurre, Jacob Piehler
Summary: Self-labeling enzymes (SLE) like the HaloTag are valuable tools in high and super-resolution fluorescence microscopy. New fluorogenic substrates for SLE allow imaging with excess dye. Two engineered variants of HaloTag7 with restored dehalogenase activity, reHaloTagS (approx. 0.006 s(-1)) and reHaloTagF (approx. 0.055 s(-1)), showed different turnover kinetics in vitro. Imaging with reHaloTag labeling demonstrated 3-5 times enhanced photostability, and single molecule imaging enabled controlled and stable labeling density over extended periods. Combination with structured illumination allowed simultaneous visualization of single molecule diffusion and organellar dynamics. These applications showcase the potential of reHaloTag labeling in advancing fluorescence microscopy techniques.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2023)
Article
Nanoscience & Nanotechnology
Ryo Yoshihara, Shuta Nomi, Kan Shoji
Summary: Scanning ion conductance microscopy (SICM) using biological nanopores is a powerful analytical tool that provides spatially resolved topography and chemical information. This study proposes a method to prepare lipid bilayers using a hydrogel-filled nanopipette, which improves the lifetime of the lipid bilayer and reduces the translocation speed of analytes through nanopores.
ACS APPLIED NANO MATERIALS
(2022)
Article
Optics
Zewei Luo, Ge Wu, Mengting Kong, Zhi Chen, Zhengfei Zhuang, Junchao Fan, Tongsheng Chen
Summary: Forster resonance energy transfer (FRET) microscopy is a valuable tool for studying biological systems, but super-resolution FRET imaging in living cells is challenging. In this study, we propose a SR FRET method called SIM-FRET that combines SR structured illumination microscopy (SIM) imaging and acceptor sensitized emission FRET imaging to achieve live-cell quantitative SR FRET imaging. Our method provides enhanced spatial resolution and maintains the advantages of quantitative FRET analysis. We validate the effectiveness of SIM-FRET through simulated models and live-cell samples, and demonstrate its ability to reveal the intricate structure of FRET signals.
PHOTONICS RESEARCH
(2023)
Review
Chemistry, Analytical
Zrinko Baricevic, Zahra Ayar, Samuel M. Leitao, Miranda Mladinic, Georg E. Fantner, Jelena Ban
Summary: Time-lapse light microscopy combined with in vitro neuronal cultures has made a significant contribution to Developmental Neuroscience. Although fluorescently labeled tags and dyes have provided advancements in cell imaging, long-term recordings remain challenging due to phototoxicity and interference with cell physiology. Hence, label-free high-resolution methods, such as scanning ion conductance microscopy (SICM) or digital holography microscopy (DHM) are proposed for studying live cell dynamics during neuronal development and regeneration.
Article
Multidisciplinary Sciences
Rajesh Ranjan, Xin Chen
Summary: This study presents a method for conducting super-resolution timelapse fluorescence live cell imaging in situ, compatible with low fluorescent intensity cells and capable of visualizing multiple subcellular structures. Special procedures in sample preparation and immobilizing specimens are necessary for this technique, but it is broadly applicable to various cell types.
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
(2021)
Article
Chemistry, Physical
Barun Kumar Maity, Duncan Nall, Yongjae Lee, Paul R. Selvin
Summary: Point Accumulation for Imaging in Nanoscale Topography (PAINT) is a single-molecule technique that achieves super-resolution microscopy with a resolution of approximately 5-25 nanometers by using exchangeable DNA oligos or peptide-pairs to create blinking phenomenon. By transfecting the protein-of-interest with a docker-coil, similar localization can be achieved at approximately 10 nm compared to using external dockers. This method allows for imaging surface proteins in live cells and provides enhanced resolution for organelles in fixed cells.
Article
Cell Biology
Vasilii Kolmogorov, Alexander Erofeev, Alexander Vaneev, Lyubov Gorbacheva, Dmitry Kolesov, Natalia Klyachko, Yuri Korchev, Petr Gorelkin
Summary: The mechanical properties of neuronal cells are crucial for various processes such as growth, generation of traction forces, adhesion, and migration. Chemical signaling disruption can lead to diseases, and glutamate excitotoxicity can affect the structural reorganization and mechanical properties of neuronal cells. Scanning ion-conductance microscopy (SICM) is a noncontact and low-stress method that can be used to investigate the mechanical properties of living neuronal cells in the presence of glutamate.
Article
Electrochemistry
Hiroki Ida, Noriko Taira, Koichi Azuma, Akichika Kumatani, Misao Akishiba, Shiroh Futaki, Yasufumi Takahashi, Hitoshi Shiku
Summary: In this study, we used scanning ion conductance microscopy to investigate the effects of five different inhibitors on membrane dynamics in living cells. Our results showed that cytochalasins and sodium azide both slowed down horizontal ruffle movements, although their inhibitory mechanisms are different. We also found that ruffle deformation and elongation were less affected compared to movements along the cell membrane direction, suggesting the involvement of mechanisms beyond actin polymerization. The disappearance of ruffles and smoothing of the cell surface were observed after treatment with 5-(N-ethyl-N-isopropyl)-Amiloride, while active ruffles without cup-like structures were observed in the presence of wortmannin. These findings contribute to a better understanding of the effects of inhibitors.
ELECTROCHIMICA ACTA
(2023)
Article
Chemistry, Multidisciplinary
Masafumi Minoshima, Taro Umeno, Kohei Kadooka, Margaux Roux, Namiko Yamada, Kazuya Kikuchi
Summary: To understand protein function in live cells, it is important to monitor protein dynamics and sense their environment in real-time. We developed a versatile chemical protein-labeling tool that efficiently forms stable carbamoylated complexes with beta-lactamase, allowing long-term visualization of labeled proteins in live cells. The use of an alpha-fluorinated carboxylate ester-based prodrug enables cell membrane permeability and stable labeling of intracellular proteins. Combining this labeling tool with a pH-activatable fluorescent probe enables visual monitoring of lysosomal protein translocation during autophagy.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2023)
Article
Chemistry, Multidisciplinary
Zheng Lv, Zhongwei Man, Hongtu Cui, Zhenzhen Xu, Huanhuan Cao, Shuai Li, Qing Liao, Qihua He, Lemin Zheng, Hongbing Fu
Summary: This study presents two efficient red-emitting aggregation-induced emission (AIE) luminogens with off-on fluorescence characteristic for specific lysosomes and mitochondria targeting. The AIE luminogens show good biocompatibility, excellent photostability, and a Stokes' shift larger than 100 nm, enabling high fidelity imaging of lysosomes and mitochondria. Moreover, dynamic lysosomal fusion and mitochondrial fission are imaged with a high resolution of 65.6 nm using stimulated emission depletion (STED) imaging. Additionally, interactions between lysosomes and mitochondria during mitophagy are recorded, providing practical guidance for designing specific organelle targeting probes for live cell dynamic super-resolution imaging.
ADVANCED FUNCTIONAL MATERIALS
(2021)
Article
Physics, Multidisciplinary
Yun-Qing Tang, Cai-Wei Zhou, Hui-Wen Hao, Yu-Jie Sun
Summary: A fully convolutional encoder-decoder network (FCEDN) was developed and applied to image scanning microscopy (ISM), achieving super-resolution imaging. The study demonstrated the feasibility and effectiveness of using this network in different microscopy systems.
Article
Chemistry, Multidisciplinary
Tom Baladi, Jesper R. Nilsson, Audrey Gallud, Emanuele Celauro, Cecile Gasse, Fabienne Levi-Acobas, Ivo Sarac, Marcel R. Hollenstein, Anders Dahlen, Elin K. Esbjorner, L. Marcus Wilhelmsson
Summary: The study introduces a stealth labeling strategy to efficiently generate RNA transcripts and translate them into correct proteins, providing an important tool for biological research and RNA delivery studies.
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
(2021)
Review
Plant Sciences
Noriyoshi Yagi, Akira Yoshinari, Ryu J. Iwatate, Reika Isoda, Wolf B. Frommer, Masayoshi Nakamura
Summary: Fluorescent probes are essential tools for visualizing cellular structures and molecular dynamics in living cells, with a wide variety of small-molecule dyes synthesized for labeling purposes. Self-labeling technologies allow these probes to be attached to cellular domains or specific proteins, and are now being increasingly utilized in plant research.
PLANT AND CELL PHYSIOLOGY
(2021)
Article
Optics
Jincheng Chen, Qiuyu Fang, Li Huang, Xin Ye, Luhong Jin, Heng Zhang, Yinqian Luo, Min Zhu, Luhao Zhang, Baohua Ji, Xiang Tian, Yingke Xu
Summary: This study developed a novel deep learning accelerated SRRF method that enables super-resolution reconstruction with only 5 low SNR images, and allows real-time visualization of microtubule dynamics and interactions with CCPs.
OPTICS AND LASERS IN ENGINEERING
(2024)
Review
Optics
Herbert Schneckenburger, Verena Richter
Summary: This short overview discusses 3D live cell imaging, relevant samples, challenges like optical sectioning and phototoxicity, and enhanced microscopy methods with some examples and applications. Future advancements may involve increasing microscopic resolution and utilizing molecular sensing techniques.
Editorial Material
Physiology
Patricia F. Lalor, Thomas Huser, Leo A. van Grunsven
FRONTIERS IN PHYSIOLOGY
(2022)
Article
Biochemical Research Methods
Dominic A. Helmerich, Gerti Beliu, Danush Taban, Mara Meub, Marcel Streit, Alexander Kuhlemann, Soeren Doose, Markus Sauer
Summary: This study demonstrates the use of time-resolved detection and photoswitching fingerprinting analysis to determine the number and distance of closely spaced fluorophores within the sub-10-nm range. The study also reveals the impact of resonance energy transfer between fluorophores on the localization probabilities of sub-10-nm fluorescence imaging.
Article
Multidisciplinary Sciences
Jan Christoph Thiele, Marvin Jungblut, Dominic A. Helmerich, Roman Tsukanov, Anna Chizhik, Alexey Chizhik, Martin J. Schnermann, Markus Sauer, Oleksii Nevskyi, Joerg Enderlein
Summary: The MIET imaging method utilizes metal-induced energy transfer to achieve nanometer-level axial resolution, and combines it with the excellent lateral resolution of dSTORM to enable three-dimensional super-resolution imaging of subcellular structures.
Article
Multidisciplinary Sciences
Alice Sandmeyer, Lili Wang, Wolfgang Huebner, Marcel Mueller, Benjamin K. Chen, Thomas Huser
Summary: Cost-effective, highly portable, and easy to use high-resolution live-cell imaging systems can revolutionize research in challenging environments, enabling real-time tracking of virus particles and studying infection mechanisms.
Article
Engineering, Biomedical
Thorsten Keller, Nora Trinks, Jessica Brand, Steffen Trippmacher, Philipp Stahlhut, Krystyna Albrecht, Georg Papastavrou, Hermann Koepsell, Markus Sauer, Juergen Groll
Summary: Nanohydrogels combine the advantages of hydrogels and nanoparticles, making them a promising drug delivery system. They can be synthesized by crosslinking polyglycidol prepolymers modified with thiol groups, resulting in disulfide bonds. This allows biomolecules like the antidiabetic peptide RS1-reg to be covalently bound to the nanogel, protecting them during gastric passage and facilitating uptake by small intestinal enterocytes.
ADVANCED HEALTHCARE MATERIALS
(2023)
Article
Microbiology
Ehsan Asghari, Bernhard Peter Kaltschmidt, Luis van Merwyk, Thomas Huser, Christian Kaltschmidt, Andreas Huetten, Barbara Kaltschmidt
Summary: Microorganisms forming a biofilm can develop multidrug resistance through information exchange. These multidrug-resistant, biofilm-producing microorganisms are responsible for many hospital-acquired infections and also cause significant damage in industrial sectors. In this study, nanoparticles of nickel and nickel oxide were found to have the highest antibacterial efficacy against Pseudomonas aeruginosa and Staphylococcus aureus isolated from household appliances. Interestingly, only P. aeruginosa was able to survive at high concentrations due to clustering toxic nanoparticles through biofilm formation.
Article
Chemistry, Physical
Anders Kokkvoll Engdahl, Oleg Grauberger, Andreas Brockhinke, Mark Schuettpelz, Thomas Huser
Summary: Photoinduced off-switching of organic fluorophores is commonly used in super-resolution microscopy for single molecule imaging. This study suggests that the interactions between sodium sulfite and glycerol can also achieve efficient switching of organic fluorophores. The addition of glycerol to the imaging buffer allows tuning of the dye switching from intermittent fluorescence to the off-state.
Article
Chemistry, Multidisciplinary
Patrick Eiring, Teresa Klein, Simone Backes, Marcel Streit, Marvin Jungblut, Soeren Doose, Gerti Beliu, Markus Sauer
Summary: By using dSTORM in combination with different labeling approaches, the study visualized and quantified the distribution and expression of ACE2 on different cells. The results showed that endogenous ACE2 receptors exist as monomers in the plasma membrane with densities of only 1-2 receptors per square micrometer. Additionally, binding of trimeric S proteins does not induce the formation of ACE2 oligomers in the plasma membrane. Infection studies demonstrated that a single S protein interaction with a monomeric ACE2 receptor is sufficient for infection, providing SARS-CoV-2 a high infectivity.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2023)
Review
Chemistry, Multidisciplinary
Gang Wen, Volker Leen, Taoufik Rohand, Markus Sauer, Johan Hofkens
Summary: Expansion microscopy (ExM) is a recently developed super-resolution technique that enables visualization of biological targets at nanoscale resolution on conventional fluorescence microscopes. This review summarizes recent advances in ExM, with a focus on the chemical aspects of the method, including chemistries for biomolecule grafting and polymer synthesis. The combination of ExM with other microscopy techniques and the impact of fixation methods on ultrastructure preservation are also discussed. The review concludes with a perspective on existing challenges and future directions.
Article
Biochemistry & Molecular Biology
Daniela Brenner, Nina Geiger, Jan Schlegel, Viktoria Diesendorf, Louise Kersting, Julian Fink, Linda Stelz, Sibylle Schneider-Schaulies, Markus Sauer, Jochen Bodem, Juergen Seibel
Summary: Recently, the effectiveness of C6-ceramides in suppressing viral replication by trapping the virus in lysosomes has been demonstrated. In this study, a synthetic ceramide derivative called AKS461 was evaluated for its antiviral activity against SARS-CoV-2. The results showed that AKS461 accumulated in lysosomes and efficiently inhibited SARS-CoV-2 replication in different cell types. This study highlights the importance of lysosomes as the central organelle for C6-ceramides to inhibit viral replication and provides a tool for studying ceramide-associated cellular and viral pathways.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2023)
Article
Chemistry, Multidisciplinary
Marvin Jungblut, Simone Backes, Marcel Streit, Georg Gasteiger, Soren Doose, Markus Sauer, Gerti Beliu
Summary: In this study, the researchers introduced a novel approach combining genetic code expansion and click chemistry to produce highly fluorescent and infectious pseudoviruses, called clickVSVs. ClickVSVs enable robust and precise virus-cell interaction studies without compromising the biological function of viral surface proteins, and have a higher efficiency in detecting surface proteins compared to classical immunolabeling.
Article
Optics
Cihang Kong, Christian Pilger, Manuel Kunisch, Christine Foerster, Jan Schulte am Esch, Thomas Huser
Summary: Coherent Raman scattering (CRS) microscopy is a label-free imaging method with high chemical sensitivity and sub-micron spatial resolution. A tunable two-color fiber laser with superior noise performance is demonstrated. Large-area multimodal imaging of human liver tissue sections is achieved and compared to commercial laser data.
LASER & PHOTONICS REVIEWS
(2023)
Article
Optics
Henning Ortkrass, Gerd Wiebusch, Jochen Linnenbruegger, Jasmin Schuerstedt, Karolina Szafranska, Peter McCourt, Thomas Huser
Summary: In this study, a novel prism telescope is introduced to seamlessly change the separation between parallel laser beams, allowing adjustment of the spatial resolution of the optical imaging system. By controlling the distance between the hexagonal prisms, the position of opposing laser spots in the back focal plane can be easily controlled, enabling a smooth transition from 2D-SIM to TIRF excitation.
Correction
Oncology
Philippa K. Brosch, Tessa Korsa, Danush Taban, Patrick Eiring, Philipp Kreisz, Sascha Hildebrand, Julia Neubauer, Heiko Zimmermann, Markus Sauer, Ryo Shirakashi, Cholpon S. Djuzenova, Dmitri Sisario, Vladimir L. Sukhorukov
Article
Chemistry, Multidisciplinary
Dominic A. Helmerich, Made Budiarta, Danush Taban, Soeren Doose, Gerti Beliu, Markus Sauer
Summary: Super-resolution microscopy has enabled direct insight into cellular structures and protein arrangements with unmatched spatial resolution. PicoRulers, designed as advanced molecular nanorulers for super-resolution fluorescence imaging, maintain structural integrity under cellular conditions and represent ideal molecular nanorulers for benchmarking the performance of super-resolution imaging techniques. The interaction among three fluorophores separated by 6 nm can be analyzed by photoswitching fingerprint analysis and their distance resolved via DNA-PAINT.
ADVANCED MATERIALS
(2023)