MINFLUX Nanoscopy: A "Brilliant" Technique Promising Major Breakthrough.

MINFLUX nanoscopy relies on the localization of single fluorophores with expected ~ 2 nm precision in 3D mapping, roughly one order of magnitude better than standard stimulated emission depletion microscopy or stochastic optical reconstruction microscopy. This "brilliant" technique takes advantage of specialized localization principles and algorithms that require only dim fluorescence signals with a minimum flux of photons; hence the name follows. With this level of performance, MINFLUX imaging and tracking should allow for the routine study of biological processes down to the molecular scale, revealing previously unresolved details in cell structures, such as the organization of calcium channels in muscle cells or the clustering of receptors in synapses.

Insights on the Mechanical Properties of SARS-CoV-2 Particles and the Effects of the Photosensitizer Hypericin.

SARS-CoV-2 is a highly pathogenic virus responsible for the COVID-19 disease. It belongs to the Coronaviridae family, characterized by a phospholipid envelope, which is crucial for viral entry and replication in host cells. Hypericin, a lipophilic, naturally occurring photosensitizer, was reported to effectively inactivate enveloped viruses, including SARS-CoV-2, upon light irradiation.

Unveiling nuclear chromatin distribution using IsoConcentraChromJ: A flourescence imaging plugin for IsoRegional and IsoVolumetric based ratios analysis.

Chromatin exhibits non-random distribution within the nucleus being arranged into discrete domains that are spatially organized throughout the nuclear space. Both the spatial distribution and structural rearrangement of chromatin domains in the nucleus depend on epigenetic modifications of DNA and/or histones and structural elements such as the nuclear envelope. These components collectively contribute to the organization and rearrangement of chromatin domains, thereby influencing genome architecture and functional regulation.

Roadmap on Label-Free Super-Resolution Imaging.

Label-free super-resolution (LFSR) imaging relies on light-scattering processes in nanoscale objects without a need for fluorescent (FL) staining required in super-resolved FL microscopy. The objectives of this Roadmap are to present a comprehensive vision of the developments, the state-of-the-art in this field, and to discuss the resolution boundaries and hurdles which need to be overcome to break the classical diffraction limit of the LFSR imaging. The scope of this Roadmap spans from the advanced interference detection techniques, where the diffraction-limited lateral resolution is combined with unsurpassed axial and temporal resolution, to techniques with true lateral super-resolution capability which are based on understanding resolution as an information science problem, on using novel structured illumination, near-field scanning, and nonlinear optics approaches, and on designing superlenses based on nanoplasmonics, metamaterials, transformation optics, and microsphere-assisted approaches.

Numerical study of transient absorption saturation in single-layer graphene for optical nanoscopy applications.

Transient absorption, or pump-probe microscopy is an absorption-based technique that can explore samples ultrafast dynamic properties and provide fluorescence-free contrast mechanisms. When applied to graphene and its derivatives, this technique exploits the graphene transient response caused by the ultrafast interband transition as the imaging contrast mechanism. The saturation of this transition is fundamental to allow for super-resolution optical far-field imaging, following the reversible saturable optical fluorescence transitions (RESOLFT) concept, although not involving fluorescence.

Toward next-generation endoscopes integrating biomimetic video systems, nonlinear optical microscopy, and deep learning.

According to the World Health Organization, the proportion of the world's population over 60 years will approximately double by 2050. This progressive increase in the elderly population will lead to a dramatic growth of age-related diseases, resulting in tremendous pressure on the sustainability of healthcare systems globally. In this context, finding more efficient ways to address cancers, a set of diseases whose incidence is correlated with age, is of utmost importance.

Image scanning microscopy with a doughnut beam: signal strength and integrated intensity.

We discuss the effects of image scanning microscopy using doughnut beam illumination on the properties of signal strength and integrated intensity. Doughnut beam illumination can give better optical sectioning and background rejection than Airy disk illumination. The outer pixels of a detector array give a signal from defocused regions, so digital processing of these (e.

Evaluation of Severe Acute Respiratory Syndrome surveillance caused by respiratory viruses in a pediatric unit, 2013 to 2019.

Objective: To evaluate severe acute respiratory syndrome surveillance in a pediatric unit.

Methods: Descriptive study of reported severe acute respiratory syndrome cases with the detection of respiratory viruses in the nasopharyngeal sample of patients hospitalized between 2013 and 2019, in a reference hospital in the Federal District, Brazil.

Results: A total of 269 children had one or more viruses detected, resulting in 280 viruses, of which 152 (54%) were respiratory syncytial virus.

Fluorescence labeling methods influence the aggregation process of α-syn differently.

In a previous study, the coexistence of different aggregation pathways of insulin and β-amyloid (Aβ) peptides was demonstrated by correlative stimulated emission depletion (STED) microscopy and atomic force microscopy (AFM). This had been explained by suboptimal proteins labeling strategies that generate heterogeneous populations of aggregating species. However, because of the limited number of proteins considered, the failure of the fluorescent labeling that occurs in a large portion of the aggregating fibrils observed for insulin and Aβ peptides, could not be considered a general phenomenon valid for all molecular systems.

On the Advent of Super-Resolution Microscopy in the Realm of Polycomb Proteins.

The genomes of metazoans are organized at multiple spatial scales, ranging from the double helix of DNA to whole chromosomes. The intermediate genomic scale of kilobases to megabases, which corresponds to the 50-300 nm spatial scale, is particularly interesting, as the 3D arrangement of chromatin is implicated in multiple regulatory mechanisms. In this context, polycomb group (PcG) proteins stand as major epigenetic modulators of chromatin function, acting prevalently as repressors of gene transcription by combining chemical modifications of target histones with physical crosslinking of distal genomic regions and phase separation.

Multiclass ultrasound-assisted extraction, clean-up and high performance liquid chromatography-tandem mass spectrometry quantification of steroid hormone residues in compost.

An analytical method for multiclass determination of steroid hormones in compost has been developed to fill the lack of methods for steroid residuals monitoring in this waste-derived product, increasingly produced and recycled in the circular-economy approach. The procedure simply entails an ultrasound-assisted extraction (UAE) on 300 mg compost by 3 × 2.5 mL methanol × 5 min sonication steps followed by a quick clean-up by solid-phase extraction (SPE) on the silica-based Supelclean™ LC-NH that avoids use of organic solvents.

SPLIT-PIN software enabling confocal and super-resolution imaging with a virtually closed pinhole.

In point-scanning microscopy, optical sectioning is achieved using a small aperture placed in front of the detector, i.e. the detection pinhole, which rejects the out-of-focus background.

From Rice Husk Ash to Silica-Supported Carbon Nanomaterials: Characterization and Analytical Application for Pre-Concentration of Steroid Hormones from Environmental Waters.

Rice husk (RH) in the rice industry is often air-burnt to obtain energy in the form of heat and RH ash (RHA) residue. In this work, RHA was applied as a starting material to obtain silica-supported carbon nanomaterials, resulting in a new reuse of a globally produced industrial waste product, in a circular economy approach. The preparation involves ultrasound-assisted one-pot oxidation with a sulfonitric mixture followed by wet oven treatment in a closed vessel.

Signal strength and integrated intensity in confocal and image scanning microscopy.

The properties of signal strength and integrated intensity in a scanned imaging system are reviewed. These properties are especially applied to confocal imaging systems, including image scanning microscopy. The integrated intensity, equal to the image of a uniform planar (sheet) object, rather than the peak of the point spread function, is a measure of the flux in an image.

Focus image scanning microscopy for sharp and gentle super-resolved microscopy.

To date, the feasibility of super-resolution microscopy for imaging live and thick samples is still limited. Stimulated emission depletion (STED) microscopy requires high-intensity illumination to achieve sub-diffraction resolution, potentially introducing photodamage to live specimens. Moreover, the out-of-focus background may degrade the signal stemming from the focal plane.

Piwil2 (Mili) sustains neurogenesis and prevents cellular senescence in the postnatal hippocampus.

Adult neural progenitor cells (aNPCs) ensure lifelong neurogenesis in the mammalian hippocampus. Proper regulation of aNPC fate has thus important implications for brain plasticity and healthy aging. Piwi proteins and the small noncoding RNAs interacting with them (piRNAs) have been proposed to control memory and anxiety, but the mechanism remains elusive.

Maximizing the Rod Outer Segment Yield in Retinas Extracted from Cattle Eyes.

The retina is a thin neuronal multilayer responsible for the detection of visual information. The first step in visual transduction occurs in the photoreceptor outer segment. The studies on photoreception and visual biochemistry have often utilized rod outer segments (OS) or OS disks purified from mammalian eyes.

Possible Association between Vogt-Koyanagi-Harada Disease and Coronavirus Disease Vaccine: A Report of Four Cases.

Vogt-Koyanagi-Harada (VKH) syndrome is an autoimmune, multisystemic disease characterized by severe bilateral granulomatous posterior, which can occur due to viral infection or vaccination. We report four cases that had a likely association between VKH disease and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination. Two patients had VKH symptoms within 1 and 2 weeks after receiving the ChAdOx1 nCoV-19 vaccine.

A red-green photochromic bacterial protein as a new contrast agent for improved photoacoustic imaging.

The GAF3 domain of the cyanobacteriochrome Slr1393 from . PCC6803, binding phycocyanobilin as a chromophore, shows photochromicity between two stable, green- and red-absorbing states, characterized by relatively high photoconversion yields. Using nanosecond-pulsed excitation by red or green light, respectively, and suitable cw photoconversion beams, we demonstrate that the light-modulatable photoacoustic waveforms arising from GAF3 can be easily distinguished from background signals originating from non-modulatable competitive absorbers and scattering media.

A photosensitizing fusion protein with targeting capabilities.

The photodynamic treatment for antimicrobial applications or anticancer therapy relies on reactive oxygen species generated by photosensitizing molecules after absorption of visible or near-infrared light. If the photosensitizing molecule is in close vicinity of the microorganism or the malignant cell, a photocytotoxic action is exerted. Therefore, the effectiveness of photosensitizing compounds strongly depends on their capability to target microbial or cancer-specific proteins.

Targeted photoimmunotherapy for cancer.

Photodynamic therapy (PDT) is a clinically approved procedure that can exert a curative action against malignant cells. The treatment implies the administration of a photoactive molecular species that, upon absorption of visible or near infrared light, sensitizes the formation of reactive oxygen species. These species are cytotoxic and lead to tumor cell death, damage vasculature, and induce inflammation.

The Interaction of Hypericin with SARS-CoV-2 Reveals a Multimodal Antiviral Activity.

Hypericin is a photosensitizing drug that is active against membrane-enveloped viruses and therefore constitutes a promising candidate for the treatment of SARS-CoV-2 infections. The antiviral efficacy of hypericin is largely determined by its affinity toward viral components and by the number of active molecules loaded on single viruses. Here we use an experimental approach to follow the interaction of hypericin with SARS-CoV-2, and we evaluate its antiviral efficacy, both in the dark and upon photoactivation.

An Efficient Green Fluorescent Protein for Stimulated Emission Depletion Super-Resolution Microscopy.

In spite of their value as genetically encodable reporters for imaging in living systems, fluorescent proteins have been used sporadically for stimulated emission depletion (STED) super-resolution imaging, owing to their moderate photophysical resistance, which does not enable reaching resolutions as high as for synthetic dyes. By a rational approach combining steady-state and ultrafast spectroscopy with gated STED imaging in living and fixed cells, we here demonstrate that F99S/M153T/V163A GFP (c3GFP) represents an efficient genetic reporter for STED, on account of no excited state absorption at depletion wavelengths <600 nm and a long emission lifetime. This makes c3GFP a valuable alternative to more common, but less photostable, EGFP and YFP/Citrine mutants for STED imaging studies targeting the green-yellow region of the optical spectrum.

Versatile Supramolecular Complex for Targeted Antimicrobial Photodynamic Inactivation.

We report the development of a supramolecular structure endowed with photosensitizing properties and targeting capability for antimicrobial photodynamic inactivation. Our synthetic strategy uses the tetrameric bacterial protein streptavidin, labeled with the photosensitizer eosin, as the main building block. Biotinylated immunoglobulin G (IgG) from human serum, known to associate with protein A, was bound to the complex streptavidin-eosin.

Phasor map analysis to investigate Hutchinson-Gilford progeria cell under polarization-resolved optical scanning microscopy.

Polarized light scanning microscopy is a non-invasive and contrast-enhancing technique to investigate anisotropic specimens and chiral organizations. However, such arrangements suffer from insensitivity to confined blend of structures at sub-diffraction level. Here for the first time, we present that the pixel-by-pixel polarization modulation converted to an image phasor approach issues an insightful view of cells to distinguish anomalous subcellular organizations.