Enhanced super-resolution microscopy by combined Airyscan and Quantum-Dot-Triexciton Imaging

ABSTRACTSuper-resolution fluorescence imaging provides critically improved information about the composition, organization and dynamics of sub-cellular structures. Quantum-Dot-Triexciton Imaging (QDTI) has been introduced as an easy-to-use sub-diffraction imaging method that achieves an almost 2-fold improvement in resolution when used with conventional confocal microscopes. Here we report an overall 3-fold increase in lateral and axial resolution compared to standard confocal microscopes by combining QDTI with the Airyscan approach.

Isotropic Three-Dimensional Dual-Color Super-Resolution Microscopy with Metal-Induced Energy Transfer

10.1101/2021.12.20.473473 ◽

2021 ◽

Author(s):

Jan Christoph Thiele ◽

Marvin Jungblut ◽

Dominic A. Helmerich ◽

Roman Tsukanov ◽

Anna Chizhik ◽

...

Keyword(s):

Energy Transfer ◽

Single Molecule ◽

Three Dimensional ◽

Super Resolution ◽

Three Dimensions ◽

Lateral Resolution ◽

Cellular Structures ◽

Axial Resolution ◽

Dual Color ◽

Over the last two decades, super-resolution microscopy has seen a tremendous development in speed and resolution, but for most of its methods, there exists a remarkable gap between lateral and axial resolution. Similar to conventional optical microscopy, the axial resolution is by a factor three to five worse than the lateral resolution. One recently developed method to close this gap is metal-induced energy transfer (MIET) imaging which achieves an axial resolution down to nanometers. It exploits the distance dependent quenching of fluorescence when a fluorescent molecule is brought close to a metal surface. In the present manuscript, we combine the extreme axial resolution of MIET imaging with the extraordinary lateral resolution of single-molecule localization microscopy, in particular with direct stochastic optical reconstruction microscopy (dSTORM). This combination allows us to achieve isotropic three-dimensional super-resolution imaging of sub-cellular structures. Moreover, we employed spectral demixing for implementing dual-color MIET-dSTORM that allows us to image and co-localize, in three dimensions, two different cellular structures simultaneously.

Fluorogenic probe for fast 3D whole-cell DNA-PAINT

10.1101/2020.04.29.066886 ◽

2020 ◽

Cited By ~ 3

Author(s):

Kenny KH Chung ◽

Zhao Zhang ◽

Phylicia Kidd ◽

Yongdeng Zhang ◽

Nathan D Williams ◽

...

Keyword(s):

3D Imaging ◽

Super Resolution ◽

Fold Increase ◽

High Fidelity ◽

Optical Sectioning ◽

High Background ◽

Fluorogenic Probe ◽

Microscopy Method ◽

Imaging Speed

AbstractDNA-PAINT is an increasingly popular super-resolution microscopy method that can acquire high-fidelity images at nanometer resolution. It suffers, however, from high background and very slow imaging speed, both of which can be attributed to the presence of unbound fluorophores in solution. We present a fluorogenic DNA-PAINT probe that solves these problems and demonstrate 3D imaging without the need for optical sectioning and a 26-fold increase in imaging speed over regular DNA-PAINT.

Single-Molecule Kinetics and Super-Resolution Microscopy by Fluorescence Imaging of Transient Binding on DNA Origami

Nano Letters ◽

10.1021/nl103427w ◽

2010 ◽

Vol 10 (11) ◽

pp. 4756-4761 ◽

Cited By ~ 399

Author(s):

Ralf Jungmann ◽

Christian Steinhauer ◽

Max Scheible ◽

Anton Kuzyk ◽

Philip Tinnefeld ◽

...

Keyword(s):

Fluorescence Imaging ◽

Single Molecule ◽

Super Resolution ◽

Dna Origami ◽

A deterministic quantum dot micropillar single photon source with >65% extraction efficiency based on fluorescence imaging method

Scientific Reports ◽

10.1038/s41598-017-13433-w ◽

2017 ◽

Vol 7 (1) ◽

Cited By ~ 9

Author(s):

Shunfa Liu ◽

Yuming Wei ◽

Rongling Su ◽

Rongbin Su ◽

Ben Ma ◽

...

Keyword(s):

Quantum Dot ◽

Fluorescence Imaging ◽

Extraction Efficiency ◽

Single Photon ◽

Imaging Method ◽

Single Photon Source ◽

Photon Source

Smoothness correction for better SOFI imaging

Scientific Reports ◽

10.1038/s41598-021-87164-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Siewert Hugelier ◽

Wim Vandenberg ◽

Tomáš Lukeš ◽

Kristin S. Grußmayer ◽

Paul H. C. Eilers ◽

...

Keyword(s):

Statistical Analysis ◽

Fluorescence Imaging ◽

Spatial Resolution ◽

Three Dimensional ◽

Super Resolution ◽

Cellular Structures ◽

Stationary Processes ◽

Two Dimensional ◽

Computationally Efficient ◽

Statistical Analysis Methods

AbstractSub-diffraction or super-resolution fluorescence imaging allows the visualization of the cellular morphology and interactions at the nanoscale. Statistical analysis methods such as super-resolution optical fluctuation imaging (SOFI) obtain an improved spatial resolution by analyzing fluorophore blinking but can be perturbed by the presence of non-stationary processes such as photodestruction or fluctuations in the illumination. In this work, we propose to use Whittaker smoothing to remove these smooth signal trends and retain only the information associated to independent blinking of the emitters, thus enhancing the SOFI signals. We find that our method works well to correct photodestruction, especially when it occurs quickly. The resulting images show a much higher contrast, strongly suppressed background and a more detailed visualization of cellular structures. Our method is parameter-free and computationally efficient, and can be readily applied on both two-dimensional and three-dimensional data.

Fluorophore-labelled RNA aptamers to common protein tags as super-resolution imaging reagents.

10.1101/2020.02.27.968578 ◽

2020 ◽

Author(s):

Juan Wang ◽

Avtar Singh ◽

Abdullah Ozer ◽

Warren R Zipfel

Keyword(s):

Fluorescent Protein ◽

Super Resolution ◽

Rna Aptamers ◽

Cellular Structures ◽

Intracellular Proteins ◽

Resolution Imaging ◽

Green Fluorescent ◽

Protein Tags ◽

Conventional Antibody

Developing labelling methods that densely and specifically label targeted cellular structures is critically important for centroid localization-based super-resolution microscopy. Being easy and inexpensive to produce in the laboratory and of relatively small size, RNA aptamers have potential as a substitute for conventional antibody labelling. By using aptamers selected against common protein tags - GFP (green fluorescent protein) in this case - we demonstrate labelling methods using dSTORM-compatible fluorophores for STORM and hybridizable imager strands for DNA-PAINT super-resolution optical imaging of any cellular proteins fused to the aptamer binding target. We show that we can label both extracellular and intracellular proteins for super-resolution imaging, and that the method in particular, offers some interesting advantages for live cell super-resolution imaging of plasma membrane proteins.

Light Microscopy of Mitochondria at the Nanoscale

Annual Review of Biophysics ◽

10.1146/annurev-biophys-121219-081550 ◽

2020 ◽

Vol 49 (1) ◽

pp. 289-308 ◽

Cited By ~ 9

Author(s):

Stefan Jakobs ◽

Till Stephan ◽

Peter Ilgen ◽

Christian Brüser

Keyword(s):

Light Microscopy ◽

Mitochondrial Function ◽

Super Resolution ◽

Diffraction Limit ◽

Cellular Structures ◽

Inner Membrane ◽

Double Membrane ◽

Future Developments ◽

Conventional Light Microscopy

Mitochondria are essential for eukaryotic life. These double-membrane organelles often form highly dynamic tubular networks interacting with many cellular structures. Their highly convoluted contiguous inner membrane compartmentalizes the organelle, which is crucial for mitochondrial function. Since the diameter of the mitochondrial tubules is generally close to the diffraction limit of light microscopy, it is often challenging, if not impossible, to visualize submitochondrial structures or protein distributions using conventional light microscopy. This renders super-resolution microscopy particularly valuable, and attractive, for studying mitochondria. Super-resolution microscopy encompasses a diverse set of approaches that extend resolution, as well as nanoscopy techniques that can even overcome the diffraction limit. In this review, we provide an overview of recent studies using super-resolution microscopy to investigate mitochondria, discuss the strengths and opportunities of the various methods in addressing specific questions in mitochondrial biology, and highlight potential future developments.

Visualizing and discovering cellular structures with super-resolution microscopy

Science ◽

10.1126/science.aau1044 ◽

2018 ◽

Vol 361 (6405) ◽

pp. 880-887 ◽

Cited By ~ 177

Author(s):

Yaron M. Sigal ◽

Ruobo Zhou ◽

Xiaowei Zhuang

Keyword(s):

State Of The Art ◽

Super Resolution ◽

Building Blocks ◽

Diffraction Limit ◽

Three Dimensions ◽

Cellular Structures ◽

Nanometer Scale ◽

Molecular Building Blocks ◽

Resolution Imaging ◽

Super-resolution microscopy has overcome a long-held resolution barrier—the diffraction limit—in light microscopy and enabled visualization of previously invisible molecular details in biological systems. Since their conception, super-resolution imaging methods have continually evolved and can now be used to image cellular structures in three dimensions, multiple colors, and living systems with nanometer-scale resolution. These methods have been applied to answer questions involving the organization, interaction, stoichiometry, and dynamics of individual molecular building blocks and their integration into functional machineries in cells and tissues. In this Review, we provide an overview of super-resolution methods, their state-of-the-art capabilities, and their constantly expanding applications to biology, with a focus on the latter. We will also describe the current technical challenges and future advances anticipated in super-resolution imaging.

Theranostics with radiation-induced ultrasound emission (TRUE)

Journal of Innovative Optical Health Sciences ◽

10.1142/s1793545818300021 ◽

2018 ◽

Vol 11 (03) ◽

pp. 1830002

Author(s):

Elijah Robertson ◽

Liangzhong Xiang

Keyword(s):

Radiation Dosimetry ◽

Imaging Techniques ◽

Super Resolution ◽

Optical Diffraction ◽

Cellular Structures ◽

Fluorescence Labeling ◽

X Ray ◽

Radiation Induced ◽

Ray Projection ◽

Two novel ultrasound imaging techniques with imaging contrast mechanisms are in the works: X-ray-induced acoustic computed tomography (XACT), and nanoscale photoacoustic tomography (nPAT). XACT has incredible potential in: (1) biomedical imaging, through which a 3D image can be generated using only a single X-ray projection, and (2) radiation dosimetry. nPAT as a new alternative of super-resolution microscopy can break through the optical diffraction limit and is capable of exploring sub-cellular structures without reliance on fluorescence labeling. We expect these new imaging techniques to find widespread applications in both pre-clinical and clinical biomedical research.

Nanocarriers used as probes for super-resolution microscopy

Materials Chemistry Frontiers ◽

10.1039/d0qm00684j ◽

2021 ◽

Author(s):

Sreejesh Sreedharan ◽

Rajeshwari Tiwari ◽

Deepak Tyde ◽

Stephen O. Aderinto ◽

Sumit Kumar Pramanik ◽

...

Keyword(s):

Single Molecule ◽

Cell Biology ◽

Super Resolution ◽

Cellular Structures ◽

Resolution Single

Super-resolution microscopy (SRM) has revolutionized cell biology, enabling visualization of cellular structures with nanometric resolution, single-molecule sensitivity, and with multiple colors. Here we review how nanocontainers have been used to enhance these techniques.