Direct optical measurement of intramolecular distances with angstrom precision DOI
Steffen J. Sahl, Jessica Matthias, Kaushik Inamdar

et al.

Science, Journal Year: 2024, Volume and Issue: 386(6718), P. 180 - 187

Published: Oct. 10, 2024

Optical investigations of nanometer distances between proteins, their subunits, or other biomolecules have been the exclusive prerogative Förster resonance energy transfer (FRET) microscopy for decades. In this work, we show that MINFLUX fluorescence nanoscopy measures intramolecular down to 1 nanometer—and in planar projections angstrom—directly, linearly, and with angstrom precision. Our method was validated by quantifying well-characterized 1- 10-nanometer polypeptides proteins. Moreover, visualized orientations immunoglobulin applied human cells, revealed specific configurations a histidine kinase PAS domain dimer. results open door examining proximities interactions direct position measurements at intramacromolecular scale.

Language: Английский

Lipid Rafts in Signalling, Diseases, and Infections: What Can Be Learned from Fluorescence Techniques? DOI Creative Commons
Sara Anselmo,

Elisa Bonaccorso,

Chiara M. A. Gangemi

et al.

Membranes, Journal Year: 2025, Volume and Issue: 15(1), P. 6 - 6

Published: Jan. 1, 2025

Lipid rafts are dynamic microdomains in the membrane, rich cholesterol and sphingolipids, that critical for biological processes like cell signalling, membrane trafficking, protein organization. Their essential role is claimed both physiological pathological conditions, including cancer, neurodegenerative diseases, viral infections, making them a key area of research. Fluorescence-based approaches, super-resolution fluorescence microscopy techniques, enable precise analysis organization, dynamics, interactions these microdomains, thanks also to innovative design appropriate fluorescent probes. Moreover, non-invasive approaches allow study live cells, facilitating collection quantitative data under physiologically relevant conditions. This review synthesizes latest insights into lipid underscores how techniques have advanced our understanding microdomains. The findings emphasize pivotal health disease, providing foundation future research potential therapeutic interventions.

Language: Английский

Citations

4
Phasor-FSTM: a new paradigm for multicolor super-resolution imaging of living cells based on fluorescence modulation and lifetime multiplexing DOI Creative Commons
Luwei Wang, Yue Chen,

Jiaqing Guo

et al.

Light Science & Applications, Journal Year: 2025, Volume and Issue: 14(1)

Published: Jan. 3, 2025

Abstract Multicolor microscopy and super-resolution optical are two widely used techniques that greatly enhance the ability to distinguish resolve structures in cellular imaging. These methods have individually transformed imaging by allowing detailed visualization of subcellular structures, as well organelle interactions. However, integrating multicolor into a single method remains challenging due issues like spectral overlap, crosstalk, photobleaching, phototoxicity, technical complexity. challenges arise from conflicting requirements using different fluorophores for labeling with specific properties We propose novel called phasor-based fluorescence spatiotemporal modulation (Phasor-FSTM). This uses time-resolved detection acquire data encoded photons, employs phasor analysis simultaneously separate multiple components, applies create images. Phasor-FSTM enables identification structural components greater spatial accuracy on an enhanced laser scanning confocal microscope single-wavelength laser. To demonstrate capabilities Phasor-FSTM, we performed two-color four-color at resolution ~λ/5 observed interactions organelles live cells during continuous duration over 20 min. Our stands out its simplicity adaptability, seamlessly fitting existing microscopes without requiring lines excitation, which also provides new avenue other technologies based principles build multi-color systems requirement lower budget.

Language: Английский

Citations

2
High-resolution and large field-of-view Fourier ptychographic microscopy and its applications in biomedicine DOI
An Pan, Chao Zuo, Baoli Yao

et al.

Reports on Progress in Physics, Journal Year: 2020, Volume and Issue: 83(9), P. 096101 - 096101

Published: July 17, 2020

Fourier ptychographic microscopy (FPM) is a promising and fast-growing computational imaging technique with high resolution, wide field-of-view (FOV) quantitative phase recovery, which effectively tackles the problems of loss, aberration-introduced artifacts, narrow depth-of-field trade-off between resolution FOV in conventional simultaneously. In this review, we provide comprehensive roadmap microscopy, fundamental principles, advantages, drawbacks existing techniques, significant roles that FPM plays development science. Since an optimization problem nature, discuss framework related work. We also reveal connection Euler's formula structured illumination microscopy. review recent advances FPM, including implementation high-precision imaging, high-throughput high-speed three-dimensional mixed-state decoupling, introduce prosperous biomedical applications. conclude by discussing challenging future can be extended to kind tackle loss system limits system. This insight used easily speckle incoherent for retina large-FOV fluorescence etc.

Language: Английский

Citations

109
Circumvention of common labelling artefacts using secondary nanobodies DOI Creative Commons
Shama Sograte‐Idrissi, Thomas Schlichthaerle, Carlos J. Duque-Afonso

et al.

Nanoscale, Journal Year: 2020, Volume and Issue: 12(18), P. 10226 - 10239

Published: Jan. 1, 2020

Secondary nanobodies can minimize probe-induced clusters artefacts. Their small size also allows fast sample penetration, and their monovalent binding enables multiplex staining using primaries from the same species.

Language: Английский

Citations

100
Chemical Analysis of Single Cells DOI
Pieter E. Oomen, Mohaddeseh Amiri-Aref, Ibrahim Kaya

et al.

Analytical Chemistry, Journal Year: 2018, Volume and Issue: 91(1), P. 588 - 621

Published: Oct. 26, 2018

ADVERTISEMENT RETURN TO ISSUEPREVReviewNEXTChemical Analysis of Single CellsPieter E. OomenPieter OomenUniversity Gothenburg, Department Chemistry and Molecular Biology, Gothenburg 41296, SwedenMore by Pieter OomenView Biographyhttp://orcid.org/0000-0001-8395-7331, Mohaddeseh A. ArefMohaddeseh ArefUniversity ArefView Biography, Ibrahim KayaIbrahim KayaUniversity SwedenDepartment Psychiatry Neurochemistry, Sahlgrenska Academy at the University Mölndal Hospital, House V3, 43180 Mölndal, SwedenThe Imaging Mass Spectrometry (Go:IMS) Laboratory, Chalmers Technology, KayaView Biographyhttp://orcid.org/0000-0003-3345-5602, Nhu T. N. PhanNhu PhanUniversity SwedenUniversity Göttingen Medical Center, Institute Neuro- Sensory Physiology, 37073, GermanyMore PhanView Andrew G. Ewing*Andrew EwingUniversity Sweden*E-mail: [email protected]More EwingView Biographyhttp://orcid.org/0000-0002-2084-0133Cite this: Anal. Chem. 2019, 91, 1, 588–621Publication Date (Web):October 26, 2018Publication History Published online26 October 2018Published inissue 2 January 2019https://pubs.acs.org/doi/10.1021/acs.analchem.8b04732https://doi.org/10.1021/acs.analchem.8b04732review-articleACS PublicationsCopyright © 2018 American Chemical SocietyRequest reuse permissionsArticle Views5585Altmetric-Citations79LEARN ABOUT THESE METRICSArticle Views are COUNTER-compliant sum full text article downloads since November 2008 (both PDF HTML) across all institutions individuals. These metrics regularly updated to reflect usage leading up last few days.Citations number other articles citing this article, calculated Crossref daily. Find more information about citation counts.The Altmetric Attention Score is a quantitative measure attention that research has received online. Clicking on donut icon will load page altmetric.com with additional details score social media presence for given article. how calculated. Share Add toView InAdd Full Text ReferenceAdd Description ExportRISCitationCitation abstractCitation referencesMore Options onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-Alertsclose SUBJECTS:Cells,Electrochemical cells,Electrodes,Ions,Microscopy Get e-Alerts

Language: Английский

Citations

91
Pulsed Interleaved MINFLUX DOI
Luciano A. Masullo, Florian Steiner, Jonas Zähringer

et al.

Nano Letters, Journal Year: 2020, Volume and Issue: 21(1), P. 840 - 846

Published: Dec. 18, 2020

We introduce p-MINFLUX, a new implementation of the highly photon-efficient single-molecule localization method with simplified experimental setup and additional fluorescence lifetime information. In contrast to original MINFLUX implementation, p-MINFLUX uses interleaved laser pulses deliver doughnut-shaped excitation foci at maximum repetition rate. Using both static dynamic DNA origami model systems, we demonstrate performance for nanoscopy tracking, respectively. delivers 1–2 nm precision 2000–1000 photon counts. addition, gives access enabling multiplexing super-resolved imaging. should help unlock full potential innovative schemes.

Language: Английский

Citations

84
Direct observation of water-mediated single-proton transport between hBN surface defects DOI
Jean Comtet, Benoît Grosjean, Evgenii Glushkov

et al.

Nature Nanotechnology, Journal Year: 2020, Volume and Issue: 15(7), P. 598 - 604

Published: May 25, 2020

Language: Английский

Citations

72
Fast widefield scan provides tunable and uniform illumination optimizing super-resolution microscopy on large fields DOI Creative Commons
Adrien Mau, Karoline Friedl, Christophe Leterrier

et al.

Nature Communications, Journal Year: 2021, Volume and Issue: 12(1)

Published: May 24, 2021

Abstract Non-uniform illumination limits quantitative analyses of fluorescence imaging techniques. In particular, single molecule localization microscopy (SMLM) relies on high irradiances, but conventional Gaussian-shaped laser restricts the usable field view to around 40 µm × µm. We present Adaptable Scanning for Tunable Excitation Regions (ASTER), a versatile technique that generates uniform and adaptable illumination. ASTER is also highly compatible with optical sectioning techniques such as total internal reflection (TIRF). For SMLM, delivers homogeneous blinking kinetics at reasonable power over fields-of-view up 200 demonstrate improves clustering analysis nanoscopic size measurements by nanorulers, microtubules clathrin-coated pits in COS-7 cells, β2-spectrin neurons. ASTER’s sharp paves way high-throughput quantification biological structures processes classical super-resolution microscopies.

Language: Английский

Citations

58
Two-photon MINFLUX with doubled localization precision DOI Creative Commons
Kun Zhao, Xinzhu Xu, Wei Ren

et al.

eLight, Journal Year: 2022, Volume and Issue: 2(1)

Published: March 25, 2022

Abstract Achieving localization with molecular precision has been of great interest for extending fluorescence microscopy to nanoscopy. MINFLUX pioneers this transition through point spread function (PSF) engineering, yet its performance is primarily limited by the signal-to-background ratio. Here we demonstrate theoretically that two-photon (2p-MINFLUX) could double PSF engineering nonlinear effect. Cramér-Rao Bound (CRB) studied as maximum precision, and CRB halved compared single-photon (1p-MINFLUX) in all three dimensions. Meanwhile, order achieve same 1p-MINFLUX, 2p-MINFLUX requires only 1/4 photons. Exploiting simultaneous excitation multiple fluorophore species, may have potential registration-free nanoscopy multicolor tracking.

Language: Английский

Citations

43
An alternative to MINFLUX that enables nanometer resolution in a confocal microscope DOI Creative Commons
Luciano A. Masullo, Alan M. Szalai, Lucía F. Lopez

et al.

Light Science & Applications, Journal Year: 2022, Volume and Issue: 11(1)

Published: June 30, 2022

Localization of single fluorescent emitters is key for physicochemical and biophysical measurements at the nanoscale beyond ensemble averaging. Examples include single-molecule tracking super-resolution imaging by localization microscopy. Among numerous methods available, MINFLUX outstands achieving a ~10-fold improvement in resolution over wide-field camera-based approaches, reaching molecular scale moderate photon counts. Widespread application related has been hindered technical complexity setups. Here, we present RASTMIN, method based on raster scanning light pattern comprising minimum intensity. RASTMIN delivers ~1-2 nm precision with usual fluorophores easily implementable standard confocal microscope few modifications. We demonstrate performance molecules DNA origami structures.

Language: Английский

Citations

42