scholarly journals Quantitative comparison of camera technologies for cost-effective Super-resolution Optical Fluctuation Imaging (SOFI)

2018 ◽  
Author(s):  
Robin Van den Eynde ◽  
Alice Sandmeyer ◽  
Wim Vandenberg ◽  
Sam Duwé ◽  
Wolfgang Hübner ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Imaging System ◽  
Fluorescent Proteins ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Cost Effective ◽  
Wide Field ◽  
Cmos Camera ◽  
Resolution Imaging ◽  
Cost Efficient

AbstractSuper-Resolution (SR) fluorescence microscopy is typically carried out on high-end research microscopes. Super-resolution Optical Fluctuation Imaging (SOFI) is a fast SR technique capable of live-cell imaging, that is compatible with many wide-field microscope systems. However, especially when employing fluorescent proteins, a key part of the imaging system is a very sensitive and well calibrated camera sensor. The substantial costs of such systems preclude many research groups from employing super-resolution imaging techniques.Here, we examine to what extent SOFI can be performed using a range of imaging hardware comprising different technologies and costs. In particular, we quantitatively compare the performance of an industry-grade CMOS camera to both state-of-the-art emCCD and sCMOS detectors, with SOFI-specific metrics. We show that SOFI data can be obtained using a cost-efficient industry-grade sensor, both on commercial and home-built microscope systems, though our analysis also readily exposes the merits of the per-pixel corrections performed in scientific cameras.

scholarly journals A spontaneously blinking fluorescent protein for simple single laser super-resolution live cell imaging

2017 ◽  
Author(s):  
Yoshiyuki Arai ◽  
Hiroki Takauchi ◽  
Yuhei Ogami ◽  
Satsuki Fujiwara ◽  
Masahiro Nakano ◽  
...  
Keyword(s):  
Single Molecule ◽  
Live Cell Imaging ◽  
Fluorescent Protein ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Light Illumination ◽  
Excitation Light ◽  
Thermally Induced ◽  
Resolution Imaging ◽  
Single Molecule Localization Microscopy

AbstractSuper-resolution imaging techniques based on single molecule localization microscopy (SMLM) broke the diffraction limit of optical microscopy in living samples with the aid of photoswitchable fluorescent probes and intricate microscopy systems. Here, we developed a fluorescent protein, SPOON, which can be switched-off by excitation light illumination and switched-on by thermally-induced dehydration resulting in an apparent spontaneous blinking behavior. This unique property of SPOON provides a simple SMLM-based super-resolution imaging platform which requires only a single 488 nm laser.

scholarly journals 3D super-resolved imaging in live cells using sub-diffractive plasmonic localization of hybrid nanopillar arrays

Nanophotonics ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 2847-2859
Author(s):  
Soojung Kim ◽  
Hyerin Song ◽  
Heesang Ahn ◽  
Seung Won Jun ◽  
Seungchul Kim ◽  
...  
Keyword(s):  
Signal To Noise Ratio ◽  
Imaging Techniques ◽  
Optical Loss ◽  
Super Resolution ◽  
Living Cells ◽  
Live Cells ◽  
Complex Biological System ◽  
Observation Volume ◽  
Resolution Imaging ◽  
Nanopillar Arrays

AbstractAnalysing dynamics of a single biomolecule using high-resolution imaging techniques has been had significant attentions to understand complex biological system. Among the many approaches, vertical nanopillar arrays in contact with the inside of cells have been reported as a one of useful imaging applications since an observation volume can be confined down to few-tens nanometre theoretically. However, the nanopillars experimentally are not able to obtain super-resolution imaging because their evanescent waves generate a high optical loss and a low signal-to-noise ratio. Also, conventional nanopillars have a limitation to yield 3D information because they do not concern field localization in z-axis. Here, we developed novel hybrid nanopillar arrays (HNPs) that consist of SiO2 nanopillars terminated with gold nanodisks, allowing extreme light localization. The electromagnetic field profiles of HNPs are obtained through simulations and imaging resolution of cell membrane and biomolecules in living cells are tested using one-photon and 3D multiphoton fluorescence microscopy, respectively. Consequently, HNPs present approximately 25 times enhanced intensity compared to controls and obtained an axial and lateral resolution of 110 and 210 nm of the intensities of fluorophores conjugated with biomolecules transported in living cells. These structures can be a great platform to analyse complex intracellular environment.

scholarly journals Comparing Super-Resolution Microscopy Techniques to Analyze Chromosomes

2021 ◽  
Vol 22 (4) ◽  
pp. 1903
Author(s):  
Ivona Kubalová ◽  
Alžběta Němečková ◽  
Klaus Weisshart ◽  
Eva Hřibová ◽  
Veit Schubert
Keyword(s):  
Single Molecule ◽  
Imaging Techniques ◽  
Chromatin Condensation ◽  
Super Resolution ◽  
Stimulated Emission ◽  
Wide Field ◽  
Structured Illumination Microscopy ◽  
Metaphase Chromosomes ◽  
Localization Microscopy ◽  
Super Resolution Microscopy

The importance of fluorescence light microscopy for understanding cellular and sub-cellular structures and functions is undeniable. However, the resolution is limited by light diffraction (~200–250 nm laterally, ~500–700 nm axially). Meanwhile, super-resolution microscopy, such as structured illumination microscopy (SIM), is being applied more and more to overcome this restriction. Instead, super-resolution by stimulated emission depletion (STED) microscopy achieving a resolution of ~50 nm laterally and ~130 nm axially has not yet frequently been applied in plant cell research due to the required specific sample preparation and stable dye staining. Single-molecule localization microscopy (SMLM) including photoactivated localization microscopy (PALM) has not yet been widely used, although this nanoscopic technique allows even the detection of single molecules. In this study, we compared protein imaging within metaphase chromosomes of barley via conventional wide-field and confocal microscopy, and the sub-diffraction methods SIM, STED, and SMLM. The chromosomes were labeled by DAPI (4′,6-diamidino-2-phenylindol), a DNA-specific dye, and with antibodies against topoisomerase IIα (Topo II), a protein important for correct chromatin condensation. Compared to the diffraction-limited methods, the combination of the three different super-resolution imaging techniques delivered tremendous additional insights into the plant chromosome architecture through the achieved increased resolution.

Design of Airborne Multi-Scale Wide-Field-of-View and High-Resolution Imaging System

2021 ◽  
Vol 41 (2) ◽  
pp. 0208002
Author(s):  
李江勇 Li Jiangyong ◽  
冯位欣 Feng Weixin ◽  
刘飞 Liu Fei ◽  
魏雅喆 Wei Yazhe ◽  
邵晓鹏 Shao Xiaopeng
Keyword(s):  
High Resolution ◽  
Imaging System ◽  
Field Of View ◽  
Wide Field ◽  
High Resolution Imaging ◽  
Multi Scale ◽  
Wide Field Of View ◽  
Resolution Imaging

Super-Resolution Imaging in Fluid Mechanics Using New Illumination Approaches

2020 ◽  
Vol 52 (1) ◽  
pp. 369-393
Author(s):  
Minami Yoda
Keyword(s):  
Fluid Mechanics ◽  
Total Internal Reflection ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Internal Reflection ◽  
Interfacial Flows ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Entire Volume ◽  
Resolution Imaging

Quantifying submillimeter flows using optical diagnostic techniques is often limited by a lack of spatial resolution and optical access. This review discusses two super-resolution imaging techniques, structured illumination microscopy and total internal reflection fluorescence or microscopy, which can visualize bulk and interfacial flows, respectively, at spatial resolutions below the classic diffraction limits. First, we discuss the theory and applications of structured illumination for optical sectioning, i.e., imaging a thin slice of a flow illuminated over its entire volume. Structured illumination can be used to visualize the interior of multiphase flows such as sprays by greatly reducing secondary scattering. Second, the theory underlying evanescent waves is introduced, followed by a review of how total internal reflection microscopy has been used to visualize interfacial flows over the last 15 years. Both techniques, which are starting to be used in fluid mechanics, could significantly improve quantitative imaging of microscale and macroscale flows.

scholarly journals Fluorescence imaging with tailored light

Nanophotonics ◽  
2019 ◽  
Vol 8 (12) ◽  
pp. 2111-2128 ◽  
Author(s):  
Jialei Tang ◽  
Jinhan Ren ◽  
Kyu Young Han
Keyword(s):  
Fluorescence Imaging ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Optical Parameters ◽  
Propagation Angle ◽  
Resolution Imaging ◽  
Time Observation ◽  
Living Organisms ◽  
Real Time Observation

AbstractFluorescence microscopy has long been a valuable tool for biological and medical imaging. Control of optical parameters such as the amplitude, phase, polarization, and propagation angle of light gives fluorescence imaging great capabilities ranging from super-resolution imaging to long-term real-time observation of living organisms. In this review, we discuss current fluorescence imaging techniques in terms of the use of tailored or structured light for the sample illumination and fluorescence detection, providing a clear overview of their working principles and capabilities.

scholarly journals Super-resolution imaging of negative-refractive graded-index photonic crystal flat lens

2021 ◽  
Author(s):  
Binming Liang ◽  
Xiao Huang ◽  
Jihong Zheng
Keyword(s):  
Photonic Crystal ◽  
Imaging System ◽  
Super Resolution ◽  
Point Sources ◽  
Image Resolution ◽  
Multiple Point ◽  
Single Image ◽  
Graded Index ◽  
Resolution Imaging ◽  
Flat Lens

Abstract Photonic crystal (PC) not only breaks through the diffraction limit of traditional lenses but also can realize super-resolution imaging. Improving the resolution is the key task of PC imaging. The main work of this paper is to use a graded-index Photonic crystal (GPC) flat lens to improve the image resolution. An air-hole type two-dimensional (2D) GPC structure based on silicon medium is proposed in this paper. Numerical simulations through RSoft reveal that when the medium in the imaging area is air, the full width at half maximum (FWHM) value of a single image reaches 0.362λ. According to the Rayleigh criterion, the images of two point sources 0.57λ apart can also be distinguished. In the imaging system composed of cedar oil and GPC flat lens, the FWHM value of a single image reaches 0.34λ. In addition, the images of multiple point sources 0.49λ apart can still be distinguished.

Super-resolution imaging of bacterial pathogens and visualization of their secreted effectors

2020 ◽  
Author(s):  
Moirangthem Kiran Singh ◽  
Linda J Kenney
Keyword(s):  
Virulence Factors ◽  
Protein Secretion ◽  
Fluorescent Probes ◽  
Imaging Techniques ◽  
Super Resolution ◽  
Bacterial Pathogenesis ◽  
Secretion Systems ◽  
Complex Protein ◽  
Resolution Imaging ◽  
Protein Secretion Systems

ABSTRACT Recent advances in super-resolution imaging techniques, together with new fluorescent probes have enhanced our understanding of bacterial pathogenesis and their interplay within the host. In this review, we provide an overview of what these techniques have taught us about the bacterial lifestyle, the nucleoid organization, its complex protein secretion systems, as well as the secreted virulence factors.

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