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| Open AccessPixel-wise programmability enables dynamic high-SNR cameras for high-speed microscopy
The researchers present an image sensor that allows every pixel to have independent exposure. It can sample a cell ROI’s fast fluorescence activity with long exposures at different phases, enhancing SNR and temporal resolution for capturing high-speed events.
- Jie Zhang
- , Jonathan Newman
- & Matthew Wilson
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Article
| Open AccessMesoscopic calcium imaging in a head-unrestrained male non-human primate using a lensless microscope
Current systems for imaging calcium dynamics in the brains of non-human primates require the animal’s movement to be restricted. Here, the authors demonstrate a mesoscale calcium imaging device in a freely moving non-human primate which features a 20 mm2 field of view.
- Jimin Wu
- , Yuzhi Chen
- & Jacob T. Robinson
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Article
| Open AccessComplexEye: a multi-lens array microscope for high-throughput embedded immune cell migration analysis
Video microscopy is key in studying cell migration, but accomplishing this in a high-throughput manner is still challenging. Here, the authors present an array microscope that can track the movements of thousands of individual cells simultaneously, and that can be used for drug screening studies.
- Zülal Cibir
- , Jacqueline Hassel
- & Matthias Gunzer
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| Open AccessThree-dimensional wide-field fluorescence microscopy for transcranial mapping of cortical microcirculation
A 3D wide-field fluorescence microscopy method is introduced based on optical astigmatism combined with fluorescence source localization. It enables transcranial cortical microcirculation mapping in murine brain with high spatiotemporal resolution.
- Quanyu Zhou
- , Zhenyue Chen
- & Daniel Razansky
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Article
| Open AccessDeep brain stimulation creates informational lesion through membrane depolarization in mouse hippocampus
The neurophysiological mechanisms of deep brain stimulation remain poorly understood. Through fluorescence voltage imaging of individual hippocampal neurons in awake mice, the authors show that deep brain stimulation causes membrane depolarization that impairs a neuron’s ability to respond to intrinsic network activity patterns and optogenetic somatic depolarization, thereby creating an informational lesion.
- Eric Lowet
- , Krishnakanth Kondabolu
- & Xue Han
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Article
| Open AccessFRET-FISH probes chromatin compaction at individual genomic loci in single cells
Chromatin compaction affects many nuclear processes yet compaction levels at individual genomic loci have been notoriously difficult to assess. Here, Ana Mota and co-authors from the Bienko-Crosetto Lab present FRET-FISH for probing chromatin compaction at selected loci in single cells.
- Ana Mota
- , Szymon Berezicki
- & Magda Bienko
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Article
| Open AccessUV photonic integrated circuits for far-field structured illumination autofluorescence microscopy
Here, the authors develop a UV-compatible photonic integrated circuit for structured illumination microscopy on a conventional wide-field microscope. Operating at a wavelength of 360 nm, they generate switchable far-field fringe patterns, and demonstrate autofluorescence imaging of yeast cells.
- Chupao Lin
- , Juan Santo Domingo Peñaranda
- & Nicolas Le Thomas
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| Open AccessAbsolute measurement of cellular activities using photochromic single-fluorophore biosensors and intermittent quantification
Biosensors often report relative rather than absolute values. Here the authors report a method that utilises the photochromic properties of biosensors to provide an absolute measure of the analyte concentration or activity: photochromism-enabled absolute quantification (PEAQ) biosensing.
- Franziska Bierbuesse
- , Anaïs C. Bourges
- & Peter Dedecker
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Article
| Open AccessAccurate localization microscopy by intrinsic aberration calibration
The authors demonstrate accurate localization in three dimensions by comprehensive calibration of an ordinary microscope, exploiting the latent information of intrinsic aberrations. Rigid transformation of the emitter positions tests the method and enables measurements in six degrees of freedom.
- Craig R. Copeland
- , Craig D. McGray
- & Samuel M. Stavis
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Article
| Open AccessFast widefield scan provides tunable and uniform illumination optimizing super-resolution microscopy on large fields
Uniform illumination is a prerequisite for quantitative analyses in both classical fluorescence microscopy and single molecule localisation microscopy. Here, the authors introduce ASTER, an illumination technique that generates uniform illumination over large and adaptable fields of view, compatible with epifluorescence, HiLo and TIRF illumination schemes.
- Adrien Mau
- , Karoline Friedl
- & Sandrine Lévêque-Fort
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Article
| Open AccessSimultaneous readout of multiple FRET pairs using photochromism
Performing multiple FRET measurements at once can be challenging. Here the authors report a method to discriminate between overlapping FRET pairs, even if the fluorophores display almost identical absorption and emission spectra, based on the photochromism of the donor fluorophores.
- Thijs Roebroek
- , Wim Vandenberg
- & Peter Dedecker
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Article
| Open AccessMetamaterial assisted illumination nanoscopy via random super-resolution speckles
Structured illumination microscopy is usually limited to 2 times spatial resolution improvement over the diffraction limit. Here, the authors introduce a metamaterial structure to generate speckle-like sub-diffraction limit illumination patterns in the near field, and achieve a 7-fold resolution improvement down to 40 nm.
- Yeon Ui Lee
- , Junxiang Zhao
- & Zhaowei Liu
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Article
| Open AccessIncorporation of sensing modalities into de novo designed fluorescence-activating proteins
Fluorescent protein reporters based on GFP exist, but have intrinsic disadvantages. Here the authors incorporate pH, Ca2+ and protein–protein interaction sensing modalities into de novo designed mini-fluorescence-activating proteins (mFAPs), with increased photostability and smaller size, which bind a range of DFHBI chromophore variants.
- Jason C. Klima
- , Lindsey A. Doyle
- & David Baker
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Article
| Open AccessSpectral cross-cumulants for multicolor super-resolved SOFI imaging
Here, the authors generalize cumulant analysis by extending it into the spectral domain to allow multicolour super-resolution optical fluctuation imaging. The simultaneous acquisition of two spectral channels followed by spectral cross-cumulant analysis and unmixing allows denser spectral and spatial sampling of the super-resolved image.
- K. S. Grußmayer
- , S. Geissbuehler
- & T. Lasser
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| Open AccessImaging of fluorescence anisotropy during photoswitching provides a simple readout for protein self-association
Performing homo-FRET measurements in cells using a fluorescence microscope is challenging, especially when using high numerical aperture objective lenses. Here the authors present a method for improved homo-FRET measurements based on anisotropy changes in photoswitchable fluorescent proteins.
- Namrata Ojha
- , Kristin H. Rainey
- & George H. Patterson
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Article
| Open AccessElectro-optic imaging enables efficient wide-field fluorescence lifetime microscopy
Nanosecond imaging techniques, such as fluorescence lifetime imaging microscopy (FLIM), are limited by low efficiency of current detectors. Here, the authors implement an electro-optic approach using Pockels cells for wide-field image gating and demonstrate high throughput FLIM on standard camera sensors.
- Adam J. Bowman
- , Brannon B. Klopfer
- & Mark A. Kasevich
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Article
| Open AccessMulti-color live-cell super-resolution volume imaging with multi-angle interference microscopy
3D super-resolution imaging of dynamic processes in live cells is still challenging, especially in a large field of view. Here the authors combine SIM with multi-angle evanescent light illumination and achieve improved lateral and axial resolution, with stack acquisition time in the range of 1–2 s.
- Youhua Chen
- , Wenjie Liu
- & Xu Liu
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| Open AccessMechanochemical feedback control of dynamin independent endocytosis modulates membrane tension in adherent cells
Plasma membrane tension is an important factor that regulates many key cellular processes. Here authors show that a specific dynamin-independent endocytic pathway is modulated by changes in tension via the mechano-transducer vinculin.
- Joseph Jose Thottacherry
- , Anita Joanna Kosmalska
- & Satyajit Mayor
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Article
| Open AccessLong-term optical brain imaging in live adult fruit flies
Time-lapse imaging studies of more than a day in the fly brain have been infeasible until now. Here the authors present a laser microsurgery approach to create a permanent window in the fly cuticle to enable time-lapse imaging of neural architecture and dynamics for up to 10–50 days.
- Cheng Huang
- , Jessica R. Maxey
- & Mark J. Schnitzer
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| Open AccessNovel genetically encoded fluorescent probes enable real-time detection of potassium in vitro and in vivo
K+ plays an important role in physiology and disease, but the lack of high specificity K+ sensors limits our understanding of its spatiotemporal dynamics. Here the authors develop genetically-encoded FRET-based probes able to quantify K+ concentration in body fluids, cells and specific organelles.
- Helmut Bischof
- , Markus Rehberg
- & Roland Malli
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Article
| Open AccessFluorescence microscopy as an alternative to electron microscopy for microscale dispersion evaluation of organic–inorganic composites
Imaging of inorganic dispersions in organic-inorganic composites is typically performed using electron microscopy. Here, the authors show that surfactants with attached aggregation-induced emission fluorophores allow simple fluorescence imaging of the spatial distribution of the inorganic filler dispersion.
- Weijiang Guan
- , Si Wang
- & Ben Zhong Tang
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Article
| Open AccessEncoding and decoding spatio-temporal information for super-resolution microscopy
Increasing the resolution of fluorescence microscopy is a fundamental need for modern cell biology. Lanzanò et al.demonstrate that arbitrary spatial resolution is, in principle, possible by encoding the fluorophore's spatial distribution information in the temporal dynamics of the fluorophore's transition.
- Luca Lanzanò
- , Iván Coto Hernández
- & Giuseppe Vicidomini
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| Open AccessLive-cell multiplane three-dimensional super-resolution optical fluctuation imaging
Super-resolution optical fluctuation imaging provides 3D images of biological specimens via blinking fluorophores. Geissbuehler et al. present a multiplexed version of this method that captures images at multiple focal planes simultaneously, reducing the acquisition time compared with standard approaches.
- Stefan Geissbuehler
- , Azat Sharipov
- & Marcel Leutenegger