Large scale superres 3D imaging: light-sheet single-molecule localization microscopy (Conference Presentation)

2017 ◽  
Author(s):  
Chieh Han Lu ◽  
Peilin Chen ◽  
Bi-Chang Chen
Keyword(s):  
Single Molecule ◽  
Large Scale ◽  
3D Imaging ◽  
Light Sheet ◽  
Localization Microscopy ◽  
Single Molecule Localization Microscopy

scholarly journals Hochaufgelöste Visualisierung einzelner Moleküle auf ganzen Zellen

BIOspektrum ◽  
2020 ◽  
Vol 26 (7) ◽  
pp. 735-738
Author(s):  
Jan Schlegel ◽  
Markus Sauer
Keyword(s):  
Single Molecule ◽  
Three Dimensional ◽  
Light Sheet ◽  
Localization Microscopy ◽  
Adhesion Receptor ◽  
Whole Cells ◽  
Analysis Techniques ◽  
Complex Interactions ◽  
Different Parts ◽  
Single Molecule Localization Microscopy

AbstractBiological systems are dynamic and three-dimensional but many techniques allow only static and two-dimensional observation of cells. We used three-dimensional (3D) lattice light-sheet single-molecule localization microscopy (dSTORM) to investigate the complex interactions and distribution of single molecules in the plasma membrane of whole cells. Different receptor densities of the adhesion receptor CD56 at different parts of the cell highlight the importance and need of three-dimensional observation and analysis techniques.

scholarly journals A real-time compression library for microscopy images

2017 ◽  
Author(s):  
Bálint Balázs ◽  
Joran Deschamps ◽  
Marvin Albert ◽  
Jonas Ries ◽  
Lars Hufnagel
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Imaging Techniques ◽  
Intrinsic Noise ◽  
Light Sheet ◽  
Localization Microscopy ◽  
Time Compression ◽  
Light Sheet Microscopy ◽  
Microscopy Images ◽  
Single Molecule Localization Microscopy

AbstractFluorescence imaging techniques such as single molecule localization microscopy, high-content screening and light-sheet microscopy are producing ever-larger datasets, which poses increasing challenges in data handling and data sharing. Here, we introduce a real-time compression library that allows for very fast (beyond 1 GB/s) compression and de-compression of microscopy datasets during acquisition. In addition to an efficient lossless mode, our algorithm also includes a lossy option, which limits pixel deviations to the intrinsic noise level of the image and yields compression ratio of up to 100-fold. We present a detailed performance analysis of the different compression modes for various biological samples and imaging modalities.

scholarly journals Oblique plane single-molecule localization microscopy for thick samples

2018 ◽  
Author(s):  
Jeongmin Kim ◽  
Michal Wojcik ◽  
Yuan Wang ◽  
Ke Xu ◽  
Xiang Zhang
Keyword(s):  
Fluorescence Detection ◽  
Single Molecule ◽  
Tissue Sample ◽  
Super Resolution ◽  
Light Sheet ◽  
Localization Microscopy ◽  
Isotropic Resolution ◽  
Fluorescent Beads ◽  
Imaging Depth ◽  
Single Molecule Localization Microscopy

We introduce single-molecule oblique plane microscopy (obSTORM) to directly image oblique sections of thick samples into depth without lengthy axial stack acquisition. Using oblique light-sheet illumination and oblique fluorescence detection, obSTORM offers uniform super-resolution throughout imaging depth in diverse biological specimens from cells to tissues. In particular, we demonstrate an isotropic resolution of ∼51 nm over a depth of 32 μm for a tissue sample, and comparable resolution over a depth of 100 μm using fluorescent beads.

Single-Molecule Imaging of Active Mitochondrial Nitroreductases using a Photo-Crosslinking Fluorescent Sensor

2019 ◽  
Author(s):  
Zacharias Thiel ◽  
Pablo Rivera-Fuentes
Keyword(s):  
Subcellular Localization ◽  
Fluorescent Probe ◽  
Single Molecule ◽  
Mammalian Cells ◽  
Fluorescent Sensor ◽  
Biological Functions ◽  
Localization Microscopy ◽  
Drug Activation ◽  
Nitroreductase Activity ◽  
Single Molecule Localization Microscopy

Many biomacromolecules are known to cluster in microdomains with specific subcellular localization. In the case of enzymes, this clustering greatly defines their biological functions. Nitroreductases are enzymes capable of reducing nitro groups to amines and play a role in detoxification and pro-drug activation. Although nitroreductase activity has been detected in mammalian cells, the subcellular localization of this activity remains incompletely characterized. Here, we report a fluorescent probe that enables super-resolved imaging of pools of nitroreductase activity within mitochondria. This probe is activated sequentially by nitroreductases and light to give a photo-crosslinked adduct of active enzymes. In combination with a general photoactivatable marker of mitochondria, we performed two-color, threedimensional, single-molecule localization microscopy. These experiments allowed us to image the sub-mitochondrial organization of microdomains of nitroreductase activity.<br>

Single-Molecule Imaging of Active Mitochondrial Nitroreductases using a Photo-Crosslinking Fluorescent Sensor

2019 ◽  
Author(s):  
Zacharias Thiel ◽  
Pablo Rivera-Fuentes
Keyword(s):  
Subcellular Localization ◽  
Fluorescent Probe ◽  
Single Molecule ◽  
Mammalian Cells ◽  
Fluorescent Sensor ◽  
Biological Functions ◽  
Localization Microscopy ◽  
Drug Activation ◽  
Nitroreductase Activity ◽  
Single Molecule Localization Microscopy

Many biomacromolecules are known to cluster in microdomains with specific subcellular localization. In the case of enzymes, this clustering greatly defines their biological functions. Nitroreductases are enzymes capable of reducing nitro groups to amines and play a role in detoxification and pro-drug activation. Although nitroreductase activity has been detected in mammalian cells, the subcellular localization of this activity remains incompletely characterized. Here, we report a fluorescent probe that enables super-resolved imaging of pools of nitroreductase activity within mitochondria. This probe is activated sequentially by nitroreductases and light to give a photo-crosslinked adduct of active enzymes. In combination with a general photoactivatable marker of mitochondria, we performed two-color, threedimensional, single-molecule localization microscopy. These experiments allowed us to image the sub-mitochondrial organization of microdomains of nitroreductase activity.<br>

scholarly journals Single-molecule localization microscopy

2021 ◽  
Vol 1 (1) ◽  
Author(s):  
Mickaël Lelek ◽  
Melina T. Gyparaki ◽  
Gerti Beliu ◽  
Florian Schueder ◽  
Juliette Griffié ◽  
...  
Keyword(s):  
Single Molecule ◽  
Localization Microscopy ◽  
Single Molecule Localization Microscopy

scholarly journals Three-dimensional total-internal reflection fluorescence nanoscopy with nanometric axial resolution by photometric localization of single molecules

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Alan M. Szalai ◽  
Bruno Siarry ◽  
Jerónimo Lukin ◽  
David J. Williamson ◽  
Nicolás Unsain ◽  
...  
Keyword(s):  
Single Molecule ◽  
Total Internal Reflection ◽  
Single Molecules ◽  
Fluorescence Microscope ◽  
Internal Reflection ◽  
Total Internal Reflection Fluorescence ◽  
Axial Position ◽  
Localization Microscopy ◽  
Localization Precision ◽  
Single Molecule Localization Microscopy

AbstractSingle-molecule localization microscopy enables far-field imaging with lateral resolution in the range of 10 to 20 nanometres, exploiting the fact that the centre position of a single-molecule’s image can be determined with much higher accuracy than the size of that image itself. However, attaining the same level of resolution in the axial (third) dimension remains challenging. Here, we present Supercritical Illumination Microscopy Photometric z-Localization with Enhanced Resolution (SIMPLER), a photometric method to decode the axial position of single molecules in a total internal reflection fluorescence microscope. SIMPLER requires no hardware modification whatsoever to a conventional total internal reflection fluorescence microscope and complements any 2D single-molecule localization microscopy method to deliver 3D images with nearly isotropic nanometric resolution. Performance examples include SIMPLER-direct stochastic optical reconstruction microscopy images of the nuclear pore complex with sub-20 nm axial localization precision and visualization of microtubule cross-sections through SIMPLER-DNA points accumulation for imaging in nanoscale topography with sub-10 nm axial localization precision.

scholarly journals Phasor based single-molecule localization microscopy in 3D (pSMLM-3D): An algorithm for MHz localization rates using standard CPUs

2018 ◽  
Vol 148 (12) ◽  
pp. 123311 ◽  
Author(s):  
Koen J. A. Martens ◽  
Arjen N. Bader ◽  
Sander Baas ◽  
Bernd Rieger ◽  
Johannes Hohlbein
Keyword(s):  
Single Molecule ◽  
Localization Microscopy ◽  
Single Molecule Localization Microscopy
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