Biological Microscopy Research Articles

PENDUGAAN UMUR CAKALANG, Katsuwonus pelamis (Linnaeus, 1758) YANG DIDARATKAN DI UPT P2SKP PONDOKDADAP SENDANGBIRU MELALUI ANALISIS LINGKARAN UMUR

Abstract Length-growth analysis of fish cannot be completed without age information. Age study on skipjack has been done through analysis of its first-hard dorsal fin. Sampling was done using 11 fish specimen taken from UPT P2SKP Pondokdadap Sendangbiru, from January to February 2018. Species identification follows the FAO standard procedure (species identification sheet) published 2001. A specimen was deposited at Depository Ichtyologicum Brawijaya with code DIB.FISH 111103. The first hard spine of 11 fish samples were cut with IsoMet low speed cutting machine at 50 mm thickness. The preparates were then analyzed using biological binocular microscope at 40x magnification. The results showed differences in number of growth annuli (annual growth-ring) at different fish length (FL). The number of annuli varied between 4 - 6+ age year at fish length (FL) variation between 38 and 62 cm. These values were closely related to those with other methods such as Modal Progression Analysis (MPA). The number of samples successfully analyzed (5 specimen) were not sufficient for analysis of length-growth parameters (L ∞ , k and t 0 ). Also, this technique did not apply for age less than yearly unit, but quite affordable (Rp.50.000,- per sample) for a fairly variable amount of specimens.

Subcellular NAMPT-mediated NAD+ salvage pathways and their roles in bioenergetics and neuronal protection after ischemic injury.

NAD+ is a cofactor required for glycolysis, tricarboxylic acid cycle, and complex I enzymatic reaction. In mammalian cells, NAD+ is predominantly synthesized through the salvage pathway, where nicotinamide phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme. Previously, we demonstrated that NAMPT exerts a neuroprotective effect in ischemia through the suppression of mitochondrial dysfunction. Mammalian cells maintain distinct NAD+ pools in the cytosol, mitochondria, and nuclei. However, it is unknown whether mitochondria have an intact machinery for NAD+ salvage, and if so, whether it plays a dominant role in bioenergetics, mitochondrial function, and neuronal protection after ischemia. Here, using mouse primary cortical neuron and cortical tissue preparations, and multiple technologies including cytosolic and mitochondrial subfractionation, viral over-expression of transgenes, molecular biology, and confocal microscopy, we provided compelling evidence that neuronal mitochondria possess an intact machinery of NAMPT-mediated NAD+ salvage pathway, and that NAMPT and nicotinamide mononucleotide adenylyltransferase 3 (NMNAT3) are localized in the mitochondrial matrix. By knocking down NMNAT1-3 and NAMPT with siRNA, we found that NMNAT3 has a larger effect on basal and ATP production-related mitochondrial respiration than NMNAT1-2 in primary cultured neurons, while NMNAT1-2 have a larger effect on glycolytic flux than NMNAT3. Using an oxygen glucose deprivation model, we found that mitochondrial, cytoplasmic, and non-subcellular compartmental over-expressions of NAMPT have a comparable effect on neuronal protection and suppression of apoptosis-inducing factor translocation. The current study provides novel insights into the roles of subcellular compartmental NAD+ salvage pathways in NAD+ homeostasis, bioenergetics, and neuronal protection in ischemic conditions.

3D sub-diffraction imaging in a conventional confocal configuration by exploiting super-linear emitters

Sub-diffraction microscopy enables bio-imaging with unprecedented clarity. However, most super-resolution methods require complex, costly purpose-built systems, involve image post-processing and struggle with sub-diffraction imaging in 3D. Here, we realize a conceptually different super-resolution approach which circumvents these limitations and enables 3D sub-diffraction imaging on conventional confocal microscopes. We refer to it as super-linear excitation-emission (SEE) microscopy, as it relies on markers with super-linear dependence of the emission on the excitation power. Super-linear markers proposed here are upconversion nanoparticles of NaYF4, doped with 20% Yb and unconventionally high 8% Tm, which are conveniently excited in the near-infrared biological window. We develop a computational framework calculating the 3D resolution for any viable scanning beam shape and excitation-emission probe profile. Imaging of colominic acid-coated upconversion nanoparticles endocytosed by neuronal cells, at resolutions twice better than the diffraction limit both in lateral and axial directions, illustrates the applicability of SEE microscopy for sub-cellular biology.

Argon Laser Peripheral Iridoplasty and Argon Laser Pupilloplasty: Alternative Management for Medically Unresponsive Acute Primary Angle Closure.

Objective To introduce the combined laser technique, argon laser peripheral iridoplasty (ALPI) and argon laser pupilloplasty (ALPP), in the management of medically unresponsive acute primary angle closure (APAC). Design Retrospective study. Methods We retrospectively reviewed the records of 23 patients (27 eyes) with APAC, who were applied ALPI and ALPP when traditional treatment failed. The visual acuity and intraocular pressure (IOP) were monitored before surgery and at 1, 2, 12, 24, and 48 h after surgery. Additionally, the angle-opening status was monitored before surgery and 48 h after the treatment by using an ultrasonic biological microscope (UBM), and the presurgical and postsurgical cornea edema statuses were observed by using a slit lamp. We also documented the complications of laser treatment. Results For the ALPI + ALPP laser-effective group, the presurgical IOP was 52.1 ± 9.3 mmHg and the postsurgical IOP was 37.6 ± 10.9 mmHg (1 h), 28.4 ± 12.4 mmHg (2 h), 19.9 ± 9.0 mmHg (6 h), 16.8 ± 7.3 mmHg (12 h), 15.9 ± 5.9 mmHg (24 h), and 14.9 ± 5.0 mmHg (48 h), with statistically significant differences (p < 0.05) in each time point. It was observed in all the patients that the corneal edema alleviated, the angles opened, and visual acuity recovered with varying degrees at 48 h after applying combined laser treatment. For the ALPI + ALPP laser-ineffective group, further interventions were taken. Definite treatment was given in both groups to maintain the long-term IOP control. Conclusions Although the combination of ALPI and ALPP is a temporizing therapeutic strategy for APAC, it is effective in relieving pupillary block which is unresponsive to miotic agents, opening the closed angle to a certain extent, restoring the transparency of cornea, and reducing IOP to a safe level for further definitive treatment.

Ratiometric BRET Measurements of ATP with a Genetically-Encoded Luminescent Sensor.

Luciferase-based reporters provide a key measurement approach in a broad range of applications, from in vitro high-throughput screening to whole animal imaging. For example, luminescence intensity is widely used to measure promoter activity, protein expression levels, and cell growth. However, luminescence intensity measurements are subject to quantitative irregularities caused by luminescence decay and variation in reporter expression level. In contrast, bioluminescence resonance energy transfer (BRET) sensors provide the advantages of luciferase-based reporters but overcome the aforementioned irregularities because of the inherently ratiometric readout. Here, we generated a new ratiometric BRET sensor of ATP (ARSeNL—ATP detection with a Ratiometric mScarlet-NanoLuc sensor), and we demonstrated that it provides a stable and robust readout across protein, cell, and whole animal tissue contexts. The ARSeNL sensor was engineered by screening a color palette of sensors utilizing variants of the high photon flux NanoLuc luciferase as donors and a panel of red fluorescent proteins as acceptors. We found that the novel combination of NanoLuc and mScarlet exhibited the largest dynamic range, with a 5-fold change in the BRET ratio upon saturation with ATP. Importantly, the NanoLuc-mScarlet BRET pair provided a large spectral separation between luminescence emission channels that is compatible with green and red filter sets extensively used in typical biological microscopes and animal imaging systems. Using this new sensor, we showed that the BRET ratio was independent of luminescence intensity decay and sensor expression level, and the BRET ratio faithfully reported differences in live-cell energy metabolism whether in culture or within mouse tissue. In particular, BRET analyte sensors have not been used broadly in tissue contexts, and thus, in principle, our sensor could provide a new tool for in vivo imaging of metabolic status.

SUN-018 LOW-VACUUM ELECTRON MICROSCOPY ANALYSIS FOR FORMALIN-FIXED PARAFFIN-EMBEDDED RAT KIDNEY SECTIONS

One of the major goal of biological microscopy is to elucidate the structural evidence relating functional activity or pathophysiological condition. High resolution imaging by electron microscopy can provide detailed tissue architectures, but the specific and skillful technics are required and it takes long time to prepare the samples. Low-vacuum scanning electron microscopy (LV-SEM) developed recently is a simple and fast technic to observe higher resolution of histological paraffin sections prepared for conventional light microscopy.

SUN-017 LOW-VACUUM SCANNING ELECTRON MICROSCOPY ANALYSIS FOR HUMAN BIOPSY SAMPLES

One of the major goal of biological microscopy is to elucidate the structural evidence relating functional activity or pathophysiological condition. High resolution imaging by electron microscopy can provide detailed tissue architectures, but the specific and skillful technics are required and it takes long time to the sample preparation. Low-vacuum scanning electron microscopy (LV-SEM) developed recently is a simple and fast technic to observe higher resolution of histological paraffin sections prepared for conventional light microscopy. Periodic acid methenamine silver (PAM) stain is a routine staining for basement membrane in renal biopsy, and contains silver, a heavy metal, which enhances the backscattered electron signal. To evaluate the benefit of LV-SEM, we observed PAM stained glomerular basement membrane (GBM) in human renal biopsy samples. The study was performed under the declaration of Helsinki principles. The study protocol was approved by the Institutional Review Board of the Tohoku Medical and Pharmaceutical University. All participants provided written informed consent after a full explanation of the purpose of the study and the potential risk involved. The human kidneys were obtained from patients with membranous nephropathy, IgA nephropathy, Alport syndrome, or thin basement membrane disease by renal biopsy and embedded in paraffin. Four-μm sections were deparaffinized with xylene, hydrated with ethanol, and stained with periodic acid methenamine silver (PAM). Miniscope TM4000 (Hitachi High-Technologies) was used to observe the GBM by detecting backscattered electron signal. After evacuation of the specimen chamber a few minutes, the sections were observed under the electron beam accelerating voltage of 10 kV for higher magnification (x2,500 to x20,000) or 15 kV for lower magnification (x40 to x2,000) with 30 Pa. The observations at higher magnification were easily performed under LV-SEM by zooming up from x40 to x20,000. The PAM-positive GBMs were bright under LV-SEM. The GBMs showed dark electron dense deposits in membranous nephropathy and IgA nephropathy, and characteristic coarse meshwork appearances in Alport syndrome. The GBMs in thin basement membrane disease seemed relatively thinner than the others. Direct observation of conventional renal biopsy paraffin sections by LV-SEM is a simple and beneficial method for the rapid and detailed analysis of the diseases. It will likely be useful for the observation of GBM morphological change.

Speckle-structured illumination for 3D phase and fluorescence computational microscopy.

High-content biological microscopy targets high-resolution imaging across large fields-of-view, often achieved by computational imaging approaches. Previously, we demonstrated 2D multimodal high-content microscopy via structured illumination microscopy (SIM) with resolution the diffraction limit, using speckle illumination from Scotch tape. In this work, we extend the method to 3D by leveraging the fact that the speckle illumination is in fact a 3D structured pattern. We use both a coherent and an incoherent imaging model to develop algorithms for joint retrieval of the 3D super-resolved fluorescent and complex-field distributions of the sample. Our reconstructed images resolve features beyond the physical diffraction-limit set by the system's objective and demonstrate 3D multimodal imaging with m3 resolution over a volume of m3.

A sequential algorithm to detect diffusion switching along intracellular particle trajectories.

Recent advances in molecular biology and fluorescence microscopy imaging have made possible the inference of the dynamics of single molecules in living cells. Changes of dynamics can occur along a trajectory. Then, an issue is to estimate the temporal change-points that is the times at which a change of dynamics occurs. The number of points in the trajectory required to detect such changes will depend on both the magnitude and type of the motion changes. Here, the number of points per trajectory is of the order of 102, even if in practice dramatic motion changes can be detected with less points. We propose a non-parametric procedure based on test statistics computed on local windows along the trajectory to detect the change-points. This algorithm controls the number of false change-point detections in the case where the trajectory is fully Brownian. We also develop a strategy for aggregating the detections obtained with different window sizes so that the window size is no longer a parameter to optimize. A Monte Carlo study is proposed to demonstrate the performances of the method and also to compare the procedure to two competitive algorithms. At the end, we illustrate the efficacy of the method on real data in 2D and 3D, depicting the motion of mRNA complexes-called mRNA-binding proteins-in neuronal dendrites, Galectin-3 endocytosis and trafficking within the cell. A user-friendly Matlab package containing examples and the code of the simulations used in the paper is available at http://serpico.rennes.inria.fr/doku.php? id=software:cpanalysis:index. Supplementary data are available at Bioinformatics online.

CM01: a facility for cryo-electron microscopy at the European Synchrotron.

Recent improvements in direct electron detectors, microscope technology and software provided the stimulus for a `quantum leap' in the application of cryo-electron microscopy in structural biology, and many national and international centres have since been created in order to exploit this. Here, a new facility for cryo-electron microscopy focused on single-particle reconstruction of biological macromolecules that has been commissioned at the European Synchrotron Radiation Facility (ESRF) is presented. The facility is operated by a consortium of institutes co-located on the European Photon and Neutron Campus and is managed in a similar fashion to a synchrotron X-ray beamline. It has been open to the ESRF structural biology user community since November 2017 and will remain open during the 2019 ESRF-EBS shutdown.

Super-Resolution Lensless Imaging of Cells Using Brownian Motion

The lensless imaging technique, which integrates a microscope into a complementary metal oxide semiconductor (CMOS) digital image sensor, has become increasingly important for the miniaturization of biological microscope and cell detection equipment. However, limited by the pixel size of the CMOS image sensor (CIS), the resolution of a cell image without optical amplification is low. This is also a key defect with the lensless imaging technique, which has been studied by a many scholars. In this manuscript, we propose a method to improve the resolution of the cell images using the Brownian motion of living cells in liquid. A two-step algorithm of motion estimation for image registration is proposed. Then, the raw holographic images are reconstructed using normalized convolution super-resolution algorithm. The result shows that the effect of the collected cell image under the lensless imaging system is close to the effect of a 10× objective lens.

Lung infection caused by Pseudomonas aeruginosa in a CD26/DPP4 deficient F344 rat model.

BackgroundPseudomonas aeruginosa (PA) is the most important opportunistic pathogen in causing nosocomial infections and, furthermore, poses a permanent threat for severe chronic infections in patients with cystic fibrosis or COPD. The transmembrane protein CD26 with dipeptidyl peptidase-4 (DPP4) activity shows an increased expression in inflamed tissue. We tested whether CD26/DPP4 deficiency leads to reduced inflammation and decreased structural damage when infected with PA.MethodsCD26/DPP4+ and CD26/DPP4− rats were instilled intratracheally with NaCl (controls) or with PA. Six hours later, bacterial distribution was detected with the in vivo imaging system 200 (IVIS). Lungs were then processed for molecular biology, light and electron microscopy and analyzed qualitatively, quantitatively and stereologically. Bacterial numbers were determined in homogenized lungs.ResultsCompared to saline treated controls, in both infected groups (1) the acinar airspace was significantly increased, (2) the volume density of the alveolar epithelium was significantly decreased, (3) the septal thickness was significantly reduced, (4) more than 40% of the alveolar epithelial surface was damaged, and up to 36% of the epithelial surface was covered with edema. In infected CD26− rats, the increase in lung weight was significantly less pronounced, the portion of edematous alveolar airspace was significantly lower and the part of edema interspersed with PA was decreased significantly.ConclusionsCD26/DPP4 deficiency resulted in reduced pulmonary edema under sublethal PA infection, implicating a role for CD26 in infection progression. The partly pronounced structural damage may mask further possible influences of CD26 on the inflammatory response.

Simulation of sub-nm carrier profiling by scanning frequency comb microscopy

A mode-locked laser focused on the tunneling junction of a scanning tunneling microscope (STM) superimposes a microwave frequency comb with hundreds of harmonics on the DC tunneling current. Each harmonic, at an integer multiple of the laser pulse repetition frequency, sets the present state-of-the-art for narrow linewidth at its frequency to enable low-noise measurements at an average laser power of several milliwatts. Measurements of the attenuation of the harmonics, which is caused by the spreading resistance, may be used to determine the resistivity of the sample. In Scanning Frequency Comb Microscopy (SFCM) feedback control of the tip-sample distance is based on the power at the harmonics. No DC bias voltage or DC tunneling current is required and the data rate is much higher than that with an STM. Simulations of the spatial distribution of the power dissipated in the sample show the feasibility of non-destructive true sub-nm resolution in the carrier profiling of semiconductors. With no DC bias voltage and no DC tunneling current band-bending and other changes to semiconductor samples in an STM are mitigated and there is a possibility for in vivo microscopy in biology and medicine.

Preparation of amphiphilic TiO2 Janus particles with highly enhanced photocatalytic activity

Stearic-acid-modified TiO2 (STA-TiO2) particles were prepared via the impregnation approach and used as a precursor for preparing TiO2 Janus particles. The morphology, structure, and properties of the TiO2 Janus particles were characterized using Fourier-transform infrared spectroscopy, ultraviolet-visible diffuse reflectance spectroscopy, thermogravimetric analysis, fluorescence microscopy, high-resolution transmission electron microscopy, contact angle analysis, dynamic light scattering, biological microscopy, X-ray photoelectron spectroscopy, and electrochemical impedance spectroscopy measurements. The results show that TiO2 Janus particles can be successfully prepared via toposelective surface modification. STA grafted on the surface of TiO2 enhances its hydrophobicity, promotes charge separation, and improves its adsorption capacity for organic compounds. The TiO2 Janus particles strongly adsorb on an oil-water interface to form a stable Pickering emulsion. The degradation rates of high-concentration kerosene and nitrobenzene wastewaters when the photocatalyst is pure TiO2, STA-TiO2, or TiO2 Janus particles are discussed and compared. The degradation rates were determined using an ultraviolet-visible spectrophotometer. It was found that the Pickering emulsion stabilized by the TiO2 Janus particles exhibited the best photocatalytic performance; these Janus particles show promising potential for catalytic application.

Neprilysin colocalizes with the V‐ATPase in kidney A‐type intercalated cells: possible role in urinary acidification

Intercalated cells (IC) in kidney collecting ducts are essential for acid‐base homeostasis. A proteomic analysis of these cells after purification by FACS revealed unexpectedly that they express high levels of neprilysin, a neutral protease, previously reported to be located in the brush border of proximal tubule cells. Therefore, we re‐examined the distribution of neprilysin in the kidney by immunohistochemistry and western blot analysis. Using anti‐neprilysin antibodies and two collecting duct markers: anti‐AQP2 antibody (principal cells) and anti‐A1 subunit v‐ATPase subunit (intercalated cells), we found that neprilysin was detectable not only in the proximal tubule, but also in ICs (but not principal cells) in all regions of the collecting duct. A similar distribution of neprilysin was seen in both mouse and rat kidneys. We then used an anti‐pendrin antibody to identify B‐type ICs in cortical collecting ducts. Only 35% of IC‐B population express neprilysin. Western blot analysis using an antibody against neprilysin showed that both cortical and medullary membrane preparations expressed neprilysin, and that female mice express more neprilysin than males in the cortex. However, this sex difference is not present in the medullary preparation, and we, therefore, attribute it mainly to a difference in proximal tubule expression. A similar result was observed in the kidneys of knockout mice that lack the IC‐specific B1 subunit of the V‐ATPase. In order to study the potential role of neprilysin in renal physiology, we treated male mice for 3 days with thiorphan, a selective neprilysin inhibitor. We measured a slight reduction of urine osmolality 1851 ± 160 to 1357 ± 240 (p=0.12) and an increase in urine volume 0.86 ± 0.21 to 1.48 ± 0.50, p= 0.23). In contrast, the change in urine pH was significant, increasing from 6.6 ± 0.1 to 6.9 ± 0.1, (n=3, p= 0.029) with the inhibitor, suggesting a role of neprilysin in acid excretion, possibly via IC function and V‐ATPase activity. In summary, we found that neprilysin is highly expressed in A‐ICs, where it colocalizes with the V‐ATPase. Its expression is higher in female mice, and it may have an as yet undetermined role in renal acid excretion.Support or Funding InformationThis work was supported by National Institutes of Health (NIH) grant DK096586 (D. Brown). The Zeiss LMS 800 confocal system in the Program in Membrane Biology (PMB) Microscopy Core was purchased using an NIH Shared Instrumentation Grant S10 RR031563‐01 (DB).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Multifunctional iron oxide-carbon hybrid nanoparticles for targeted fluorescent/MR dual-modal imaging and detection of breast cancer cells

An efficient method for the highly sensitive and specific detection of cancer cells is crucial for the early diagnosis of cancer. In this work, we propose a one-pot approach to fabricating magnetic-fluorescent iron oxide-carbon hybrid nanomaterials (MCNP) with excellent stable, high quantum yield and excellent magnetic properties for breast cancer cells recognition and detection via magnetic resonance and multicolour fluorescence imaging. MCNPs were efficiently synthesised via one-pot, multi-component reactions of FeCl3, FeCl2•4H2O, citric acid and ethylenediamine in diethylene glycol. The MCNPs showed strong excitation wavelength-dependent fluorescence in the blue-red region with a high quantum yield of 58.4%, and they presented higher stability and T2 relaxivity than pure iron oxide nanoparticles. After conjugating with CD44 monoclonal antibodies, the fabricated targeting nanoprobe, MCNPs-CD44, demonstrated a specific fluorescence/MRI dual imaging contrast effect in 4T1 breast cancer cells. Biological transmission electron microscope imaging showed a significant preferential uptake of the nanoparticle conjugates by the 4T1 cells. By taking advantage of the high binding affinity and specificity of the CD44 antibodies to the overexpressed CD44 on the cancer cell surface, the developed MCNPs-CD44 probe distinguished 4T1 breast cancer cells from normal cells and detected as low as a few hundred cancer cells, thus indicating the potential application of multifunctional nanocomposites in the MR diagnosis and fluorescence positioning of breast cancer at cellular-level resolution.

Quantitative STEM imaging of electron beam induced mass loss of epoxy resin sections

In sample preparation of biological samples for electron microscopy, many types of embedding media are widely used. Unfortunately, none of them is perfectly resistant to beam induced damage. The article is focused on mass loss measuring of pure epoxy resin EMbed 812 that replaced Epon – the most widely used embedding resin for biological electron microscopy, in a form of ultrathin sections with thicknesses ranging from 30 to 100 nm. The STEM imaging was performed in a quantitative way which allowed us to estimate the mass loss directly up to the total dose of 3000 e−/nm2. For data acquisition we used SEM equipped with a commercial STEM detector working at a relatively low acceleration voltage of 30 kV. In this study we estimated the influence of various factors which can affect the endurance of the epoxy resin EMbed 812 ultrathin sections under an electron beam, such as the sample aging, differences between storing the samples in forms of ultrathin sections and whole blocks, ultrathin sections thicknesses, temperature of the sample, probe current, and one or two-sided carbon coating of ultrathin sections. The aim of this work is to investigate beam induced mass loss at electron energies of SEM and find out how to reduce the mass loss.

Ultrasonographic features of intraocular lymphoma

Objective To describe the ultrasonographic features of intraocular lymphoma. Methods It was a retrospective analysis of 19 patients (26 eyes) with intraocular lymphoma at the department of ophthalmology, Beijing Tongren Hospital from January 2015 to December 2017. The diagnosis was established by tissue biopsy in 4 patients with diagnostic vitrectomy and 8 with enucleation, and other 7 cases were clinically diagnosed followed by pathological examination of central nervous system lymphoma. The ultrasonographic features of all patients were analyzed. Results Of the 19 patients (26 eyes) with intraocular lymphoma, 12 cases (19 eyes) of vitreoretinal lymphoma accounted for 63.2%, of which 7 cases were bilateral eyes, and 7 cases (7 eyes) of uveal lymphoma accounted for 36.8%, all of which were monocular. Ultrasonography of vitreoretinal lymphoma included vitreous opacity, posterior vitreous detachment, partial retinal detachment, irregular hypoechoic lesions under the retina, and CDFI detected blood flow signals in the lesion. There were two different types of ultrasound findings in uveal lymphoma. Five cases (5 eyes) showed diffuse thickening of the uveal membrane with low echo, and irregular hypoechoic mass around the optic nerve and the ball wall, in which CDFI detected abundant blood flow signals. Ultrasound biological microscope examination showed that the iris-ciliary body thickened and the internal echo decreased. This ultrasound finding accounted for 71.4% of uveal lymphoma. Two cases (2 eyes) showed a solitary ciliary-choroid choroidal hypoechoic mass, combined with retinal detachment and blood flow signals with CDFI, accounting for 28.6% of uveal lymphoma. Conclusions The ultrasonography of intraocular lymphoma is various. The ultrasound findings of vitreoretinal lymphoma are not specific, while the diffuse thickened uvea and retrobulbar hypoechoic lesions encircled optic nerve and eye wall suggest the possibility of uveal lymphoma. Key words: Ultrasonography, Doppler, color; Uvea diseases; Intraocular lymphoma

Computational structured illumination for high-content fluorescence and phase microscopy.

High-content biological microscopy targets high-resolution imaging across large fields-of-view (FOVs). Recent works have demonstrated that computational imaging can provide efficient solutions for high-content microscopy. Here, we use speckle structured illumination microscopy (SIM) as a robust and cost-effective solution for high-content fluorescence microscopy with simultaneous high-content quantitative phase (QP). This multi-modal compatibility is essential for studies requiring cross-correlative biological analysis. Our method uses laterally-translated Scotch tape to generate high-resolution speckle illumination patterns across a large FOV. Custom optimization algorithms then jointly reconstruct the sample's super-resolution fluorescent (incoherent) and QP (coherent) distributions, while digitally correcting for system imperfections such as unknown speckle illumination patterns, system aberrations and pattern translations. Beyond previous linear SIM works, we achieve resolution gains of 4× the objective's diffraction-limited native resolution, resulting in 700 nm fluorescence and 1.2 μm QP resolution, across a FOV of , giving a space-bandwidth product (SBP) of 60 megapixels.

Video-rate large-scale imaging with Multi-Z confocal microscopy.

Fast, volumetric imaging over large scales has been a long-standing challenge in biological microscopy. To address this challenge, we report an augmented variant of confocal microscopy that uses a series of reflecting pinholes axially distributed in the detection space, such that each pinhole probes a different depth within the sample. We thus obtain simultaneous multiplane imaging without the need for axial scanning. Our microscope technique is versatile and configured here to provide two-color fluorescence imaging with a field of view larger than a millimeter at video rate. Its general applicability is demonstrated with neuronal imaging of both Caenorhabditis elegans and mouse brains in vivo.