Quantitatively mapping local quality of super-resolution microscopy by rolling Fourier ring correlation

Light Science & Applications, Journal Year: 2023, Volume and Issue: 12(1)

Published: Dec. 14, 2023

In fluorescence microscopy, computational algorithms have been developed to suppress noise, enhance contrast, and even enable super-resolution (SR). However, the local quality of images may vary on multiple scales, these differences can lead misconceptions. Current mapping methods fail finely estimate quality, challenging associate SR scale content. Here, we develop a rolling Fourier ring correlation (rFRC) method evaluate reconstruction uncertainties down scale. To visually pinpoint regions with low reliability, filtered rFRC is combined modified resolution-scaled error map (RSM), offering comprehensive concise for further examination. We demonstrate their performances various imaging modalities, resulting quantitative maps better integrated from different reconstructions. Overall, expect that our framework become routinely used tool biologists in assessing image datasets general inspire advances rapidly developing field imaging.

Language: Английский

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Spatiotemporal resolution in high-speed atomic force microscopy for studying biological macromolecules in action

Microscopy, Journal Year: 2023, Volume and Issue: 72(2), P. 151 - 161

Published: Feb. 6, 2023

Abstract High-speed atomic force microscopy (HS-AFM) is a unique approach that allows direct real-time visualization of biological macromolecules in action under near-physiological conditions, without any chemical labeling. Typically, the temporal resolution sub-100 ms, and spatial 2–3 nm lateral direction ∼0.1 vertical direction. A wide range biomolecular systems their dynamic processes have been studied by HS-AFM, providing deep mechanistic insights into how biomolecules function. However, level detail gleaned from an HS-AFM experiment critically depends on spatiotemporal system. In this review article, we explain principle describe determined. We also discuss recent attempts to improve further extend observable phenomena.

Language: Английский

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Does nano basic building-block of C-S-H exist? – A review of direct morphological observations

Materials & Design, Journal Year: 2024, Volume and Issue: 238, P. 112699 - 112699

Published: Feb. 1, 2024

Despite significant advancements in microstructural characterization methods, the interconnections between nanostructure and morphological diversity of calcium-silicate-hydrate (C-S-H), primary binding phase modern concrete, remain unclear. This review delves into state-of-the-art experimental findings morphology C-S-H comprehensively analyses various influencing factors. The focus here is to address long-standing debate: whether there are fundamental structural units, either fractural globule or nano sheet, that assemble form diverse microstructures. We critically assess formation involves structured assembly layered with an approximate size 4–10 nm, rather than occurring randomly. Such may have substantial heterogeneity deformability, often blurring distinction globular sheet models. Finally, this paper offers perspectives on future research directions aimed at further unravelling intricate structure C-S-H.

Language: Английский

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Label-Free Techniques for Probing Biomolecular Condensates

ACS Nano, Journal Year: 2024, Volume and Issue: 18(16), P. 10738 - 10757

Published: April 12, 2024

Biomolecular condensates play important roles in a wide array of fundamental biological processes, such as cellular compartmentalization, regulation, and other biochemical reactions. Since their discovery first observations, an extensive expansive library tools has been developed to investigate various aspects properties, encompassing structural compositional information, material evolution throughout the life cycle from formation eventual dissolution. This Review presents overview expanded set methods that researchers use probe properties biomolecular across diverse scales length, concentration, stiffness, time. In particular, we review recent years' exciting development label-free techniques methodologies. We broadly organize into 3 categories: (1) imaging-based techniques, transmitted-light microscopy (TLM) Brillouin (BM), (2) force spectroscopy atomic (AFM) optical tweezer (OT), (3) microfluidic platforms emerging technologies. point out tools' key opportunities, challenges, future perspectives analyze correlative potential well compatibility with techniques. Additionally, namely, differential dynamic (DDM) interferometric scattering (iSCAT), have huge for applications studying condensates. Finally, highlight how some these can be translated diagnostics therapy purposes. hope this serves useful guide new field aids advancing biophysical study

Language: Английский

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Cobalt nanoparticles intercalation coupled with tellurium-doping MXene for efficient electrocatalytic water splitting

Journal of Colloid and Interface Science, Journal Year: 2024, Volume and Issue: 675, P. 379 - 390

Published: July 4, 2024

Language: Английский

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Structural basis of pH-dependent activation in a CLC transporter

Nature Structural & Molecular Biology, Journal Year: 2024, Volume and Issue: 31(4), P. 644 - 656

Published: Jan. 26, 2024

Language: Английский

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Retention mechanism in slalom chromatography: Perspectives on the characterization of large DNA and RNA biopolymers in cell and gene therapy

Journal of Chromatography A, Journal Year: 2025, Volume and Issue: 1743, P. 465691 - 465691

Published: Jan. 23, 2025

Significant progress has been made in the last two decades producing small (<2μm), high-purity, and low-adsorption particles, columns system hardware, for ultra-high pressure liquid chromatography (UHPLC). Simultaneously, recent rapid expansion of cell gene therapies treating diseases necessitates novel analytical technologies analyzing large (>2 kbp) plasmid double-stranded (ds) DNA (which encodes vitro transcription (IVT) single-stranded (ss) mRNA therapeutics) dsRNAs (related to IVT production impurities) biopolymers. In this context, slalom (SC), a retention mode co-discovered 1988, is being revitalized using most advanced column improved determination critical quality attributes (CQAs) such new therapeutics. review, we first recall non-exhaustively main currently available techniques (enzyme-linked immunosorbent assay (ELISA), agarose gel electrophoresis (AGE), pulse field (PFGE), capillary (CGE), mass photometry (MP), anion-exchange (AEX), ion-pairing reversed-phase (IP-RPLC), hydrophobic interaction (HIC), size-exclusion (SEC), hydrodynamic (HDC), highly converging flow ultra-filtration (HCF-UF), asymmetrical field-flow fractionation (AF4), spectrometry (MS), atomic force microscopy (AFM)) mixtures containing nucleic acid biopolymers, while assessing their strengths weaknesses. We then focus comprehensively on SC technique, report its past applications since birth, review detail history evolution proposed mechanisms accounting observations SC. This includes emphasizes latest physico-chemical insights (shear rates packed HPLC columns, entropic elasticity relaxation dsDNA, dsRNA, biopolymers) governing behavior biopolymers Finally, based advancements understanding fundamentals SC, provide some perspectives proof-of-concept characterization by dsDNAs (plasmid digests, polymerase chain reaction (PCR) verification), separation supercoiled/circular linear linearization), isolation quantification impurities present samples produced IVT, differentiation between dsRNA conformers.

Language: Английский

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A structural biology compatible file format for atomic force microscopy

Nature Communications, Journal Year: 2025, Volume and Issue: 16(1)

Published: Feb. 15, 2025

Cryogenic electron microscopy (cryo-EM), X-ray crystallography, and nuclear magnetic resonance (NMR) contribute structural data that are interchangeable, cross-verifiable, visualizable on common platforms, making them powerful tools for our understanding of protein structures. Unfortunately, atomic force (AFM) has so far failed to interface with these biology methods, despite the recent development localization AFM (LAFM) allows extracting high-resolution information from data. Here, we build LAFM develop a pipeline transforms into 3D-density files (.afm) readable by programs commonly used visualize, analyze, interpret We show 3D-LAFM densities can serve as fields steer molecular dynamics flexible fitting (MDFF) obtain models previously unresolved states based observations in close-to-native environment. Besides, .afm format enables direct 3D or 2D visualization analysis conventional images. anticipate file will find wide usage embed repertoire methods routinely community, allowing researchers deposit repositories comparison cross-verification other techniques.

Language: Английский

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Membrane-mediated protein interactions drive membrane protein organization

Nature Communications, Journal Year: 2022, Volume and Issue: 13(1)

Published: Nov. 30, 2022

The plasma membrane's main constituents, i.e., phospholipids and membrane proteins, are known to be organized in lipid-protein functional domains supercomplexes. No active membrane-intrinsic process is establish organization. Thus, the interplay of thermal fluctuations biophysical determinants membrane-mediated protein interactions must considered understand Here, we used high-speed atomic force microscopy kinetic elastic theory investigate behavior a model oligomerization assembly controlled lipid environments. We find that hydrophobic mismatch modulates energetics, 2D Our experimental theoretical frameworks reveal how organization can emerge from Brownian diffusion minimal set physical properties constituents.

Language: Английский

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Structural Identification of Individual Helical Amyloid Filaments by Integration of Cryo-Electron Microscopy-Derived Maps in Comparative Morphometric Atomic Force Microscopy Image Analysis

Journal of Molecular Biology, Journal Year: 2022, Volume and Issue: 434(7), P. 167466 - 167466

Published: Jan. 22, 2022

The presence of amyloid fibrils is a hallmark more than 50 human disorders, including neurodegenerative diseases and systemic amyloidoses. A key unresolved challenge in understanding the involvement disease to explain relationship between individual structural polymorphs fibrils, potentially mixed populations, specific pathologies with which they are associated. Although cryo-electron microscopy (cryo-EM) solid-state nuclear magnetic resonance (ssNMR) spectroscopy methods have been successfully employed recent years determine structures high resolution detail, rely on ensemble averaging fibril entire sample or significant subpopulations. Here, we report method for identification imaged by atomic force (AFM) integration high-resolution maps determined cryo-EM comparative AFM image analysis. This approach was demonstrated using hitherto structurally formed vitro from fragment tau (297–391), termed ‘dGAE’. Our established unequivocally that dGAE bear no heparin-induced vitro. Furthermore, our analysis resulted prediction closely related paired helical filaments (PHFs) isolated Alzheimer’s (AD) brain tissue characterised cryo-EM. These results show utility particle AFM, provide workflow how data can be incorporated into facilitate an integrated polymorphism.

Language: Английский

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