Salinity is a global issue limiting efficient agricultural production. Nano-enabled plant salt tolerance is a hot topic. However, the role of nanoparticles induced possible early stimulation on ...antioxidant system in its improved plant salt tolerance is still largely unknown. Here, poly (acrylic) acid coated nanoceria (cerium oxide nanoparticles) (PNC, 7.8 nm, −31 mV) with potent ROS (reactive oxygen species) scavenging ability are used. Compared with control, no significant difference of H2O2 and O2•─ content, MDA (malondialdehyde) content, relative electric conductivity, and Fv/Fm was found in leaves and/or roots of cucumber before onset of salinity stress, regardless of leaf or root application of PNC. While, before onset of salinity stress, compared with control, the activities of SOD (superoxide dismutase, up to 1.8 folds change), POD (peroxidase, up to 2.5 folds change) and CAT (catalase, up to 2.3 folds change), and the content of GSH (glutathione, up to 3.0 folds change) and ASA (ascorbic acid, up to 2.4 folds change) in leaves and roots of cucumber with PNC leaf spray or root application were significantly increased. RNA seq analysis further confirmed that PNC foliar spray upregulates more genes in leaves over roots than the root application. These results showed that foliar sprayed PNC have stronger early stimulation effect on antioxidant system than the root applied one and leaf are more sensitive to PNC stimulation than root. After salt stress, cucumber plants with foliar sprayed PNC showed better improvement in salt tolerance than the root applied one. Also, plants with foliar sprayed PNC showed significant higher whole plant cerium content than the root applied one after salt stress. In summary, we showed that foliar spray of nanoceria is more optimal than root application in terms of improving cucumber salt tolerance, and this improvement is associated with better stimulation on antioxidant system in plants.
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•PNC foliar spray showed better uptake of Ce in cucumber than the root application.•Foliar sprayed PNC enabled better cucumber salt tolerance than its root application.•Stimulation on antioxidant system is linked with PNC enabled cucumber salt tolerance.
Out-of-plane ferroelectricity with a high transition temperature in ultrathin films is important for the exploration of new domain physics and scaling down of memory devices. However, depolarizing ...electrostatic fields and interfacial chemical bonds can destroy this long-range polar order at two-dimensional (2D) limit. Here we report the experimental discovery of the locking between out-of-plane dipoles and in-plane lattice asymmetry in atomically thin In_{2}Se_{3} crystals, a new stabilization mechanism leading to our observation of intrinsic 2D out-of-plane ferroelectricity. Through second harmonic generation spectroscopy and piezoresponse force microscopy, we found switching of out-of-plane electric polarization requires a flip of nonlinear optical polarization that corresponds to the inversion of in-plane lattice orientation. The polar order shows a very high transition temperature (∼700 K) without the assistance of extrinsic screening. This finding of intrinsic 2D ferroelectricity resulting from dipole locking opens up possibilities to explore 2D multiferroic physics and develop ultrahigh density memory devices.
Carbon nanomaterials have been widely explored in bioimaging and biosensing, but their application in forensic science and technology is rare. In this report, we show one-pot synthesis of cationic ...Carbon Dots (cCDs) and their application in fingerprinting analysis. The cCDs show narrow size distribution (<10 nm), decent quantum yield (c.a. 46%), and a band gap of 2.75 eV. The cCDs exhibit cationic charge and show excitation-dependent multi-fluorescence characteristics. The cCDs show no production of reactive oxygen species and thus are safe to use. Interestingly, they also possess bright liquid as well as solid-state fluorescence. We demonstrate that a well-resolved patterned fingerprint can be obtained within a short time (2–3 min) from different physical surfaces and can be studied using a simple cell phone camera enabling accurate visualization up to third-degree detailing of the fingerprint. Owing to multi-fluorescence characteristics of cCDs, a detailed investigation of the fluorescent patterns was carried out by a three-dimensional multiphoton confocal microscopy. In comparison to traditional methods, the cCDs fluorescence-based fingerprinting shows improvements towards rapid image capture and analysis. Thus, the cCDs form a promising material for rapid fingerprint detection in forensic sciences.
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Good sense: A highly sensitive and selective ratiometric fluorescent probe for Fe2+ was developed by linking an Fe2+‐insensitive fluorophore to an Fe2+‐sensitive reporter–fluorophore fragment. The ...fluorescence intensity ratio increased linearly with the concentration of Fe2+ (see scheme). The probe was applied to the ratiometric imaging of intracellular and adscititious fluctuations of Fe2+ in live HL‐7702 and HepG2 cells.
Translocator Protein 18 kDa (TSPO), previously named Peripheral Benzodiazepine Receptor, is a well-validated and widely used biomarker of neuroinflammation to assess diverse central nervous system ...(CNS) pathologies in preclinical and clinical studies. Many studies have shown that in animal models of human neurological and neurodegenerative disease and in the human condition, TSPO levels increase in the brain neuropil, and this increase is driven by infiltration of peripheral inflammatory cells and activation of glial cells. Therefore, a clear understanding of the dynamics of the cellular sources of the TSPO response is critically important in the interpretation of Positron Emission Tomography (PET) studies and for understanding the pathophysiology of CNS diseases.
Within the normal brain compartment, there are tissues and cells such as the choroid plexus, ependymal cells of the lining of the ventricles, and vascular endothelial cells that also express TSPO at even higher levels than in glial cells. However, there is a paucity of knowledge if these cell types respond and increase TSPO in the diseased brain. These cells do provide a background signal that needs to be accounted for in TSPO-PET imaging studies. More recently, there are reports that TSPO may be expressed in neurons of the adult brain and TSPO expression may be increased by neuronal activity. Therefore, it is essential to study this topic with a great deal of detail, methodological rigor, and rule out alternative interpretations and imaging artifacts.
High levels of TSPO are present in the outer mitochondrial membrane. Recent studies have provided evidence of its localization in other cellular compartments including the plasma membrane and perinuclear regions which may define functions that are different from that in mitochondria. A greater understanding of the TSPO subcellular localization in glial cells and infiltrating peripheral immune cells and associated function(s) may provide an additional layer of information to the understanding of TSPO neurobiology. This review is an effort to outline recent advances in understanding the cellular sources and subcellular localization of TSPO in brain cells and to examine remaining questions that require rigorous investigation.
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•A pyrazolo1,5-a pyrimidine (PyPP)-based probe for reversible detection of histidine/HSA-BSA.•It clearly distinguishes histidine from other amino acids, including cysteine.•PyPP ...exhibited low cytotoxicity and good cell permeability.•Transient PyPP-Ni2+ complex was successfully applied to detect histidine and histidine-rich proteins at a cellular level.
Amino acids and proteins are ubiquitous in all biological processes. Therefore, fluctuations in their levels profoundly impact the general well-being of any human being. Structural similarities between amino acids hamper their direct detection by fluorophores. Thus, it becomes crucial to employ an ancillary approach to accomplish this goal. In this work, the selective fluorescence response of Ni2+ towards a probe PyPP was used as a precursor for selective histidine sensing. The probe showed selectivity towards Ni2+ by fluorescence quenching, and it could be revived only in the presence of histidine. The limit of detection (LOD) for Ni2+ was 4 x10-6 M, whereas LOD for histidine was 1.08 × 10−6 M. The Ni2+ mediated selectivity could also be extended to histidine-rich proteins bovine serum albumin (BSA) and human serum albumin (HSA). The devised system was also applied to a macrophage cell line (RAW 264.7) to indicate the presence of Ni2+/histidine/HSA-BSA via confocal fluorescence microscopy, and quantitative response could be studied with flow cytometry.
•We successfully demonstrated the effectiveness and efficiency of an innovative deep learning-based approach (DLM) for automatically detecting and classifying local Ca2+ release events exemplified in ...the context of cardiomyocytes.•The DLM can, be adapted and applied to all full-frame confocal data and datasets representing Ca2+ signals. Previously analyzed data could be used to train DLM datasets collected from other cell types and in different contexts.•The suggested methodology allows researchers to detect local Ca2+ release events automatically while simultaneously classifying them in a few minutes without human intervention and bias, enabling more efficient data processing than previous methods.•The DLM approach can detect more than 75 % of the Ca²⁺ release events independently of the noise characteristics in the original recordings.•The robustness and accuracy of the DLM was demonstrated in comparison to conventional full-frame confocal imaging analysis of local intracellular Ca2+ events, as well as in comparison with the result of manual analysis by experts on site, which was previously considered the gold standard for the classification of Ca2+ events.
The release of Ca2+ ions from intracellular stores plays a crucial role in many cellular processes, acting as a secondary messenger in various cell types, including cardiomyocytes, smooth muscle cells, hepatocytes, and many others. Detecting and classifying associated local Ca2+ release events is particularly important, as these events provide insight into the mechanisms, interplay, and interdependencies of local Ca2+release events underlying global intracellular Ca2+signaling. However, time-consuming and labor-intensive procedures often complicate analysis, especially with low signal-to-noise ratio imaging data.
Here, we present an innovative deep learning-based approach for automatically detecting and classifying local Ca2+ release events. This approach is exemplified with rapid full-frame confocal imaging data recorded in isolated cardiomyocytes.
To demonstrate the robustness and accuracy of our method, we first use conventional evaluation methods by comparing the intersection between manual annotations and the segmentation of Ca2+ release events provided by the deep learning method, as well as the annotated and recognized instances of individual events. In addition to these methods, we compare the performance of the proposed model with the annotation of six experts in the field. Our model can recognize more than 75 % of the annotated Ca2+ release events and correctly classify more than 75 %. A key result was that there were no significant differences between the annotations produced by human experts and the result of the proposed deep learning model.
We conclude that the proposed approach is a robust and time-saving alternative to conventional full-frame confocal imaging analysis of local intracellular Ca2+ events.
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In recent years, the zebrafish has become a well-established laboratory model. We describe here the ZeBraInspector (ZBI) platform for high-content 3D imaging (HCI) of 5 days post-fertilization ...zebrafish eleuthero-embryos (EEs). This platform includes a mounting method based on 3D-printed stamps to create a grid of wells in an agarose cast, facilitating batch acquisitions with a fast-confocal laser scanning microscope. We describe reference labeling in cleared fish with a fluorescent lipophilic dye. Based on this labeling, the ZBI software registers. EE 3D images, making it possible to visualize numerous identically oriented EEs on a single screen, and to compare their morphologies and any fluorescent patterns at a glance. High-resolution 2D snapshots can be extracted. ZBI software is therefore useful for diverse high-content analyses (HCAs). Following automated segmentation of the lipophilic dye signal, the ZBI software performs volumetric analyses on whole EEs and their nervous system white matter. Through two examples, we illustrate the power of these analyses for obtaining statistically significant results from a small number of samples: the characterization of a phenotype associated with a neurodevelopmental mutation, and of the defects caused by treatments with a toxic anti-cancer compound.
A workflow for 3D‐CLEM investigating liver tissue KREMER, A.; VAN HAMME, E.; BONNARDEL, J. ...
Journal of microscopy (Oxford),
March 2021, 2021-03-00, 20210301, Letnik:
281, Številka:
3
Journal Article
Recenzirano
Summary
Correlative light and electron microscopy (CLEM) is a method used to investigate the exact same region in both light and electron microscopy (EM) in order to add ultrastructural information ...to a light microscopic (usually fluorescent) signal. Workflows combining optical or fluorescent data with electron microscopic images are complex, hence there is a need to communicate detailed protocols and share tips & tricks for successful application of these methods. With the development of volume‐EM techniques such as serial blockface scanning electron microscopy (SBF‐SEM) and Focussed Ion Beam‐SEM, correlation in three dimensions has become more efficient. Volume electron microscopy allows automated acquisition of serial section imaging data that can be reconstructed in three dimensions (3D) to provide a detailed, geometrically accurate view of cellular ultrastructure. In addition, combining volume‐EM with high‐resolution light microscopy (LM) techniques decreases the resolution gap between LM and EM, making retracing of a region of interest and eventual overlays more straightforward. Here, we present a workflow for 3D CLEM on mouse liver, combining high‐resolution confocal microscopy with SBF‐SEM. In this workflow, we have made use of two types of landmarks: (1) near infrared laser branding marks to find back the region imaged in LM in the electron microscope and (2) landmarks present in the tissue but independent of the cell or structure of interest to make overlay images of LM and EM data. Using this approach, we were able to make accurate 3D‐CLEM overlays of liver tissue and correlate the fluorescent signal to the ultrastructural detail provided by the electron microscope. This workflow can be adapted for other dense cellular tissues and thus act as a guide for other three‐dimensional correlative studies.
Lay Description
As cells and tissues exist in three dimensions, microscopy techniques have been developed to image samples, in 3D, at the highest possible detail. In light microscopy, fluorescent probes are used to identify specific proteins or structures either in live samples, (providing dynamic information), or in fixed slices of tissue. A disadvantage of fluorescence microscopy is that only the labeled proteins/structures are visible, while their cellular context remains hidden. Electron microscopy is able to image biological samples at high resolution and has the advantage that all structures in the tissue are visible at nanometer (10−9 m) resolution. Disadvantages of this technique are that it is more difficult to label a single structure and that the samples must be imaged under high vacuum, so biological samples need to be fixed and embedded in a plastic resin to stay as close to their natural state as possible inside the microscope. Correlative Light and Electron Microscopy aims to combine the advantages of both light and electron microscopy on the same sample. This results in datasets where fluorescent labels can be combined with the high‐resolution contextual information provided by the electron microscope. In this study we present a workflow to guide a tissue sample from the light microscope to the electron microscope and image the ultra‐structure of a specific cell type in the liver. In particular we focus on the incorporation of fiducial markers during the sample preparation to help navigate through the tissue in 3D in both microscopes. One sample is followed throughout the workflow to visualize the important steps in the process, showing the final result; a dataset combining fluorescent labels with ultra‐structural detail.
Excitons in atomically thin transition-metal dichalcogenides (TMDs) have been established as an attractive platform to explore polaritonic physics, owing to their enormous binding energies and giant ...oscillator strength. Basic spectral features of exciton polaritons in TMD microcavities, thus far, were conventionally explained via two-coupled-oscillator models. This ignores, however, the impact of phonons on the polariton energy structure. Here we establish and quantify the threefold coupling between excitons, cavity photons, and phonons. For this purpose, we employ energy-momentum-resolved photoluminescence and spatially resolved coherent two-dimensional spectroscopy to investigate the spectral properties of a high-quality-factor microcavity with an embedded WSe_{2} van der Waals heterostructure at room temperature. Our approach reveals a rich multibranch structure which thus far has not been captured in previous experiments. Simulation of the data reveals hybridized exciton-photon-phonon states, providing new physical insight into the exciton polariton system based on layered TMDs.
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