Long-term super-resolution imaging of mitochondrial dynamics

Monitoring dynamics of mitochondria has become an essential approach to explore the function of mitochondria in living cells with the emergence of super-resolution fluorescence microscopy. However, long-term super-resolution imaging of mitochondria is still challenging due to the lack of photostable fluorescent probes and stable mitochondria-specific markers which are not affected by the changes of mitochondrial membrane potential. Here, we introduce a method for long-term imaging mitochondrial dynamic through the SNAP-tag fluorogenic probe based on 4-azetidinyl-naphthalimide derivatives. Using structured illumination microscopy (SIM), we observed the fusion and fission of mitochondria over a course of 16 min at 109 nm resolution. Furthermore, the interactions as well as fusion between mitochondria and lysosomes were studied during mitophagy at the nanoscale. Convincingly, the combination of SNAP-tag fluorogenic probes and super-resolution fluorescence microscopy will offer a new way to monitor dynamic mitochondria in living cells.     

Donor-Acceptor Typed AIE Luminogens with Near-infrared Emission for Super-resolution Imaging

Aggregation-induced emission(AIE)luminogens(AIEgens)with high brightness in aggregates exhibit great potentials in biological imaging,but these AIEgens are seldom applied in super-resolution biological imaging,especially in the imaging by using the structural illumination microscope(SIM).Based on this consideration,we synthesized the donor-acceptor typed AIEgen of DTPA-BTN,which not only owns high brightness in the near-infrared(NIR)emission region from 600 nm to 1000 nm(photoluminescence quantum yield,PLQYs=11.35%),but also displays excellent photo-stability.In addition,AIE nanoparticles based on 4,7-ditriphenylamine-[1,2,5]-thiadiazolo[3,4-c]pyridine(DTPA-BTN)were also prepared with highly emissive features and excellent biocompatibility.Finally,the developed DTPA-BTN-based AIE nanoparticles were applied in the super-resolution cellular imaging via SIM,where much smaller full width at half-maximum values and high signal to noise ratios were obtained,indicating the superior imaging resolution.The results here imply that highly emissive AIEgens or AIE nanoparticles can be promising imaging agents for super-resolution imaging via SIM.     

Super-resolution in time-reversal focusing on a moving source

This paper presents a detailed analysis of time-reversal experiments involving a moving point source that emits a pulse. Different configurations are addressed with full-aperture or partial-aperture time-reversal mirrors and with subsonic or supersonic sources. Doppler effects and lack of source-receiver reciprocity significantly affect the time-reversal refocusing when the velocity of the source becomes comparable as or larger than the speed of propagation. The main result is that refocusing can be enhanced when the velocity of the source becomes close to the speed of propagation compared to the classical diffraction-limited refocusing properties when the source does not move, and this super-resolution effect can be quantified by simple and explicit formulas.     

Long-term super-resolution imaging of mitochondrial dynamics

Monitoring dynamics of mitochondria has become an essential approach to explore the function of mitochondria in living cells with the emergence of super-resolution fluorescence microscopy. However, long-term super-resolution imaging of mitochondria is still challenging due to the lack of photostable fluorescent probes and stable mitochondria-specific markers which are not affected by the changes of mitochondrial membrane potential. Here, we introduce a method for long-term imaging mitochondrial dynamic through the SNAP-tag fluorogenic probe based on 4-azetidinyl-naphthalimide derivatives. Using structured illumination microscopy (SIM), we observed the fusion and fission of mitochondria over a course of 16 min at 109 nm resolution. Furthermore, the interactions as well as fusion between mitochondria and lysosomes were studied during mitophagy at the nanoscale. Convincingly, the combination of SNAP-tag fluorogenic probes and super-resolution fluorescence microscopy will offer a new way to monitor dynamic mitochondria in living cells.     

Microstructure, optical and static recording properties of AgOx film, and near-field simulation of sub-wavelength aperture based on the AgOx marks’ formation

Structures, spectra and surface topographies of as-deposited and annealed AgO_x films have been investigated by an X-ray diffractometer, a spectrophotometer and an atomic force microscopy (AFM). X-ray diffraction and spectrum results show that the as-deposited AgO_x films with high oxygen ratios (x≥0.5) are in amorphous states and Ag crystalline particles will separate out after annealed. AFM results show that the film surface will become much rougher and film thickness will increase greatly after annealed due to the decomposition of AgO_x with release of oxygen. Static recording results show that two microstructures of the recording marks can be produced: one is the bubble mark at a low recording power and the other is the rupture bubble with an ablated aperture (hole) in the center at a high recording power. Based on the formation of rupture bubble marks, the near-field optical distribution of a focused Gaussian laser beam through a sub-wavelength aperture (200 nm in diameter) has been simulated using finite-difference-time-domain (FDTD) method. Results show that the spot size can be greatly squeezed with still highly transmitted intensity, which may lead to the super-resolution readout.     

Infrared chemical imaging: Spatial resolution evaluation and super-resolution concept

Chemical imaging systems help to solve many challenges in various scientific fields. Able to deliver rapid spatial and chemical information, modern infrared spectrometers using Focal Plane Array detectors (FPA) are of great interest. Considering conventional infrared spectrometers with a single element detector, we can consider that the diffraction-limited spatial resolution is more or less equal to the wavelength of the light (i.e. 2.5-25 μm). Unfortunately, the spatial resolution of FPA spectroscopic setup is even lower due to the detector pixel size. This becomes a real constraint when micron-sized samples are analysed. New chemometrics methods are thus of great interest to overcome such resolution drawback, while keeping our far-field infrared imaging spectrometers. The aim of the present work is to evaluate the super-resolution concept in order to increase the spatial resolution of infrared imaging spectrometers using FPA detectors. The main idea of super-resolution is the fusion of several low-resolution images of the same sample to obtain a higher-resolution image. Applying the super-resolution concept on a relatively low number of FPA acquisitions, it was possible to observe a 30% decrease in spatial resolution.     

Experimental investigation of the quality of lensless super-resolution ghost imaging via sparsity constraints

Ghost imaging via sparsity constraints (GISC) can nonlocally realize super-resolution imaging. Factors influencing the quality of lensless super-resolution GISC are investigated and the experimental results show that, the quality of GISC is enhanced as the object?s sparse ratio in the representation basis or the spatial transverse coherence lengths on the object plane are decreased. The differences between ghost imaging (GI) and GISC are also discussed.     

An adaptive super-resolution method based on regional pixel information and ringing artifacts suppression

Multi-frame image super-resolution (SR) aims to utilize information from a set of low-resolution (LR) images to compose a high-resolution (HR) one. In this paper, a novel multi-frame image super-resolution algorithm is proposed based on regional pixel information and ringing artifacts suppression. Firstly, a new regularization term which adopts Regional Adaptive Weight Coefficients (RAWC) is produced to keep edges and flat regions. After detailed analysis, an iterative process is given for image reconstruction. Then an adaptive term according to the local variance of iterative correction image is designed to evaluate the ringing artifacts. Finally, the original iteration is updated by adding the restraint term for better visual effects and lower noise of reconstructive HR image. Thorough experimental results show the proposed algorithm is effective for SR reconstruction and ringing artifacts suppression.     

FDTD study of subwavelength imaging by a photonic crystal slab

The problem of subwavelength imaging via a photonic crystal slab lens made of two-dimensional (2D) square arrays of parallel dielectric cylinders in air is studied and discussed theoretically. The finite-difference time-domain method is employed to investigate the unique features of imaging by such lens. We confirm earlier findings that a photonic crystal slab lens can provide the imaging of a point source. By analysing the transmission properties of the proposed structure, we demonstrate that inside the all-angle negative refraction, there are some favourable directions for waves to travel. We show that the surface termination of the photonic crystal is a key parameter to obtain a good quality image. The super-resolution of two sources separated by a distance less than the wavelength is also considered. It is shown that the achievable resolution is limited by the slab length.     

Simultaneous two color image capture for sub-diffraction localization fluorescence microscopy

A sub-diffraction limit fluorescence localization microscope was constructed using a standard cooled 1.4 mega-pixel fluorescence charge-coupled device (CCD) camera to simultaneously resolve closely adjacent paired quantum dots on a flat surface with emissions of 540 and 630 nm. The images of the overlapping Airy discs were analyzed to determine the center of the point spread function after noise reduction using Fourier transformation analysis. The Cartesian coordinates of the centers of the point spread functions were compared in serial images. Histograms constructed from serial images fit well to Gaussian functions for resolving two quantum dots separated by as little as 10 nm in the x–y coordinates. Statistical analysis of multiple pairs validated discrimination of inter-fluorophore distances that vary by 10 nm. The method is simple and developed for x–y resolution of dilute fluorophores on a flat surface, not serial z sectioning.