- 标题
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Development of ZnO Nanoparticles as an Efficient Zn Fertilizer: Using Synchrotron-Based Techniques and Laser Ablation to Examine Elemental Distribution in Wheat Grain
摘要： Zinc (Zn) deficiency is an important problem worldwide, adversely impacting human health. Using a field trial in China, we compared the foliar-application of both ZnO nanoparticles (ZnO-NPs) and ZnSO4 on winter wheat (Triticum aestivum L.) for increasing the Zn concentration within the grain. We also used synchrotron-based X-ray fluorescence microscopy (μ-XRF) and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) to examine the distribution of Zn within the grain. We found that ZnO-NPs increase the Zn concentration in the wheat grain, increasing from 18 mg·kg-1 in the control up to 40 mg·kg-1 when the ZnO-NPs were applied four times. These grain Zn concentrations in the ZnO-NP-treated grains are similar to those recommended for human consumption. However, the ZnO-NPs were similar in their effectiveness to ZnSO4. When examining trace element distribution in the grain, the trace elements were found to accumulate primarily in the aleurone layer and the crease region across all treatments. Importantly, Zn concentrations in the grain endosperm increased by nearly 30-fold relative to the control, with this markedly increasing Zn within the edible portion. These results demonstrate that ZnO-NPs is a suitable fertilizer for increasing Zn within wheat grain and can potentially be used to improve human nutrition.
关键词： X-ray fluorescence microscopy (μ-XRF),foliar application,wheat,Zn biofortification,laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS),ZnO-NPs
更新于2025-09-19 17:13:59
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Laser switching contrast microscopy to monitor free and restricted diffusion inside the cell nucleus
摘要： A novel microscopic technique termed laser switching contrast microscopy (LSCM) allows for the imaging of the dynamics of optically switchable proteins in single cell compartments. We present an application for the monitoring of diffusive properties of single molecules of the photo-switchable fluorescent protein Dreiklang (DRK). LSCM in the cell nucleus of Chinese hamster ovary (CHO) cells cytoplasmically expressing DRK unravels quick diffusive equilibration of the DRK molecules inside the whole cytoplasm and inside the cell nucleus within seconds. The nuclear membrane is also highly permeable for DRK. Inside the nucleus entirely distinct regions are found that only partially enable diffusive protein redistribution with mean square displacement proportional to time while in other regions the mobility of the proteins seems to be restricted. After photo-switching string like patterns of light DRK molecules are observed in the cell nucleus. In addition a fraction of these DRK molecules appears immobile. The findings support recent theories of the cell interior described as a random obstacle model with an additional immobile fraction of DRK. Numerical simulations show that at different illumination intensity and different distance from the laser focus similar patterns for fluorescence recovery might be obtained in spite of strongly varying diffusion constants.
关键词： random walk,Fluorescence microscopy,diffusion constant,nanofluidics,Dreiklang,superresolution microscopy,fluorescent protein,photo-switchable molecules
更新于2025-09-16 10:30:52
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Fast objective coupled planar illumination microscopy
摘要： Among optical imaging techniques light sheet fluorescence microscopy is one of the most attractive for capturing high-speed biological dynamics unfolding in three dimensions. The technique is potentially millions of times faster than point-scanning techniques such as two-photon microscopy. However light sheet microscopes are limited by volume scanning rate and/or camera speed. We present speed-optimized Objective Coupled Planar Illumination (OCPI) microscopy, a fast light sheet technique that avoids compromising image quality or photon efficiency. Our fast scan system supports 40 Hz imaging of 700 μm-thick volumes if camera speed is sufficient. We also address the camera speed limitation by introducing Distributed Planar Imaging (DPI), a scaleable technique that parallelizes image acquisition across cameras. Finally, we demonstrate fast calcium imaging of the larval zebrafish brain and find a heartbeat-induced artifact, removable when the imaging rate exceeds 15 Hz. These advances extend the reach of fluorescence microscopy for monitoring fast processes in large volumes.
关键词： Distributed Planar Imaging,calcium imaging,light sheet fluorescence microscopy,zebrafish brain,OCPI microscopy
更新于2025-09-16 10:30:52
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[IEEE 2019 Conference on Lasers and Electro-Optics Europe & European Quantum Electronics Conference (CLEO/Europe-EQEC) - Munich, Germany (2019.6.23-2019.6.27)] 2019 Conference on Lasers and Electro-Optics Europe & European Quantum Electronics Conference (CLEO/Europe-EQEC) - Investigation of Photoluminescence Properties of 2D-Nanoparticles to be Used as Fluorescence Markers in Microscopy
摘要： Transition metal dichalcogenides (TMDs) are direct bandgap semiconductors [1], and they exhibit fascinating optical properties, such as valley polarization, room temperature excitons, and high optical nonlinearity. Single and few-layer TMDs and TMD-nanoparticles exhibit much stronger photoluminescence than the bulk material [2]. This property makes them an attractive candidate for use as mesoscale fluorescent markers, e.g., for use in fluorescence microscopy. As opposed to conventional fluorescent markers, such as fluorescent molecules they do not exhibit phototoxicity and have been shown to be biocompatible in contrast to, e.g., many types of quantum dots. Here we focus on the characterization of MoS2 and WS2 nanoparticles, obtained through liquid phase exfoliation [3], from powdered precursors. Sonication of TMD powder in N-Methyl pyrrolidone (NMP) is carried out, as NMP and single layer TMDs have comparable surface energies to prevent the restacking of exfoliated nanosheets. The resulting solution was found to have particles of a typical size of 65 nm, which have been investigated for PL-spectra and long-term PL-behaviour, such as blinking and bleaching. Structural analysis reveals (See Fig.1 (a)) that we have a mixture of particles and particle agglomerates. PL spectra show that both exhibit comparable spectral signatures with peak wavelengths (Fig. 1(b)) ranging from 600 to 640nm. The excitation wavelength was 532nm. We find that these results indicate, that TMD-nanoparticles are suitable, e.g., for use as fluorescent guide stars [4] in adaptive optical microscopy in inhomogeneous samples, e.g., tissue. 2D-nanoparticles would be infiltrated into the sample, and the PL-emission could be used by a combination of a wavefront-sensor and an adaptive optical element to undo the effects of scattering in the vicinity of the particle. This method would leverage the strong PL, biocompatibility and long-term stability of the 2D-nanoparticles, to allow for longer observation cycles in sensitive samples, such as living tissue.
关键词： nanoparticles,photoluminescence,biocompatibility,fluorescence microscopy,Transition metal dichalcogenides
更新于2025-09-16 10:30:52
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Hybrid Imaging of Aspergillus fumigatus Pulmonary Infection with Fluorescent, 68Ga-Labelled Siderophores
摘要： Aspergillus fumigatus (A. fumigatus) is a human pathogen causing severe invasive fungal infections, lacking sensitive and selective diagnostic tools. A. fumigatus secretes the siderophore desferri‐triacetylfusarinine C (TAFC) to acquire iron from the human host. TAFC can be labelled with gallium‐68 to perform positron emission tomography (PET/CT) scans. Here, we aimed to chemically modify TAFC with fluorescent dyes to combine PET/CT with optical imaging for hybrid imaging applications. Starting from ferric diacetylfusarinine C ([Fe]DAFC), different fluorescent dyes were conjugated (Cy5, SulfoCy5, SulfoCy7, IRDye 800CW, ATTO700) and labelled with gallium‐68 for in vitro and in vivo characterisation. Uptake assays, growth assays and live‐cell imaging as well as biodistribution, PET/CT and ex vivo optical imaging in an infection model was performed. Novel fluorophore conjugates were recognized by the fungal TAFC transporter MirB and could be utilized as iron source. Fluorescence microscopy showed partial accumulation into hyphae. μPET/CT scans of an invasive pulmonary aspergillosis (IPA) rat model revealed diverse biodistribution patterns for each fluorophore. [68Ga]Ga‐DAFC‐Cy5/SufloCy7 and ‐IRDye 800CW lead to a visualization of the infected region of the lung. Optical imaging of ex vivo lungs corresponded to PET images with high contrast of infection versus non‐infected areas. Although fluorophores had a decisive influence on targeting and pharmacokinetics, these siderophores have potential as a hybrid imaging compounds combining PET/CT with optical imaging applications.
关键词： fluorescence microscopy,PET,near infrared,siderophores,gallium‐68,invasive pulmonary aspergillosis
更新于2025-09-16 10:30:52
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Scanning acoustic microscopy of quantum dot aggregates
摘要： Quantum dot (QD) aggregate formation is essential, especially, in nanoparticle-based drug delivery systems. Imaging of these targeted QD aggregates exposes information about the the treatment of many diseases including cancer. Scanning Acoustic Microscopy (SAM) is a non-invasive and rapid imaging modality, which can obtain qualitative and quantitative features simultaneously. In our study, acoustic impedance microscopy of QD aggregates was performed by SAM for evaluating the potential of SAM in the detection of lead-sulphide (PbS), graphene and cadmium-telluride/cadmium sulphide (CdTe/CdS) quantum dot aggregates for the first time. Absorption spectra of QDs, obtained with an ultraviolet-visible spectrometer, and fluorescence spectra of QD aggregates, obtained with an inverted fluorescence microscope, are also demonstrated. The success of imaging quantum dot aggregates by SAM indicated the potential of SAM in monitoring the micro-environment of the disease and also the therapeutic effect of the drug-loaded QD aggregates.
关键词： scanning acoustic microscopy,quantum dot aggregates,fluorescence microscopy
更新于2025-09-12 10:27:22
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Simplified Dynabeads Method Using Light Microscopy for Enumerating TCD4+ Lymphocytes in Resource-Limited Settings
摘要： Background: We demonstrated feasibility of implanting the Dynabeads method for CD4+ T lymphocyte enumeration in resource-poor settings (ANRS 1226 study). However, as this technique requires a fluorescence microscope which is not usually available in these settings, WHO has encouraged to simplify the method allowing TCD4+ lymphocyte counting under a light microscope. Methods: TCD4+ lymphocytes enumeration was assessed using Dynabeads after staining cells nuclei with non-fluorescent dyes and readings under light microscope (DLM). A total of 305 triple of values of CD4 cells counts were generated by both Dynabeads method using a light microscopy (DLM), Dynabeads method using a fluorescent microscope (DFM) and the single-platform flow cytometry technique (FCM). The accuracy of DLM was analyzed using 4 fresh blood samples showing 200, 400, 500 and 1000 cells/μl in FCM respectively. Correlations have been studied between the 3 methods. The DLM was then evaluated for its ability to correctly segregate absolute TCD4+ lymphocyte values at the thresholds of 200 cells/μl and 350 cells/μl. Findings: Cells nuclei staining with Sternheimer-Malbin, Turck1, and Giemsa allows TCD4+ lymphocytes enumeration using DLM. FCM has shown the greatest standard deviations and amplitudes. The reproducibility of DLM was better than FCM. The correlation coefficient between FCM and DFM was 0.975 and it was 0.973, 0.972 and 0.969 with DLM using Sternheimer-Malbin, Turck1 and Giemsa, respectively. The ability of DLM to correctly segregate TCD4+ lymphocyte values at the threshold of 200 cells/μl and 350 cells/μl was good. Conclusion: Reliable TCD4+ enumeration can be obtained with DLM. These results will contribute in resource-limited-settings to further reduce the cost of TCD4+ lymphocytes counting and make it more widely available in peripheral laboratories and even in central laboratories that face problems with maintenance and stock-out of reagents for flow cytometers.
关键词： Dynabeads method,Flow Cytometry,Resource-limited settings,Fluorescence microscopy,Light microscopy,TCD4+ lymphocytes enumeration
更新于2025-09-11 14:15:04
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[Methods in Molecular Biology] Barley Volume 1900 (Methods and Protocols) || Preparation of Barley Roots for Histological, Structural, and Immunolocalization Studies Using Light and Electron Microscopy
摘要： Microscopic investigations of biological objects are an integral part in plant research and most ?elds of life sciences. They allow the description of morphological, histological, and structural aspects of individual cells or tissues. Based on various cell biological tools and methods it is possible to characterize different plant genotypes or study their adaptation to changing environmental conditions. In combination with antibodies raised against speci?c antigens and epitopes light and electron microscopy enable investigation of the function of single genes/proteins in plant growth and development or their role related to abiotic or biotic stresses. Here, we describe sample preparation of barley roots for cell biological investigations using light and electron microscopy, to characterize morphological, structural, and functional aspects on root sections and the root surface.
关键词： Light microscopy,Electron microscopy,Fluorescence microscopy,Immunolocalization,Histology,Immunogold labeling
更新于2025-09-10 09:29:36
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Quenched Stochastic Optical Reconstruction Microscopy (qSTORM) with Graphene Oxide
摘要： Quenched Stochastic Optical Reconstruction Microscopy (qSTORM) was demonstrated with graphene oxide sheets, peptides and bacteria; a method of contrast enhancement with super-resolution fluorescence microscopy. Individual sheets of graphene oxide (GO) were imaged with a resolution of 16 nm using the quenching of fluorescence emission by GO via its large Resonant Energy Transfer (RET) efficiency. The method was then extended to image self-assembled peptide aggregates (resolution 19 nm) and live bacterial cells (resolution 55 nm, the capsular structure of E. coli from urinary tract infections) with extremely low backgrounds and high contrasts (between one and two orders of magnitude contrast factor improvements that depended on the thickness of the graphene oxide layer used). Graphene oxide films combined with STORM imaging thus provide an extremely convenient method to image samples with large backgrounds due to non-specifically bound fluorophores (either due to excess labelling or autofluorescent molecules), which is a common occurrence in studies of both biological cells and soft-condensed matter. The GO quenches the fluorescence across a thin layer at distances of less than 15 nm. Graphene oxide films coated with thin layers (≤15 nm) of polystyrene, polymethylmethacrylate and polylysine are shown to be effective in producing high contrast qSTORM images, providing a convenient modulation of sample/substrate interactions. The GO coatings can also provide an increased image resolution and a factor of 2.3 improvement was observed with the peptide fibres using a feature of interest metric,when there was a large non-specifically bound background.
关键词： Quenched Stochastic Optical Reconstruction Microscopy,contrast enhancement,peptide aggregates,qSTORM,fluorescence quenching,RET,bacterial cells,Resonant Energy Transfer,graphene oxide,super-resolution fluorescence microscopy
更新于2025-09-10 09:29:36
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Correlative 3D x-ray fluorescence and ptychographic tomography of frozen-hydrated green algae
摘要： Accurate knowledge of elemental distributions within biological organisms is critical for understanding their cellular roles. The ability to couple this knowledge with overall cellular architecture in three dimensions (3D) deepens our understanding of cellular chemistry. Using a whole, frozen-hydrated Chlamydomonas reinhardtii cell as an example, we report the development of 3D correlative microscopy through a combination of simultaneous cryogenic x-ray ptychography and x-ray fluorescence microscopy. By taking advantage of a recently developed tomographic reconstruction algorithm, termed GENeralized Fourier Iterative REconstruction (GENFIRE), we produce high-quality 3D maps of the unlabeled alga’s cellular ultrastructure and elemental distributions within the cell. We demonstrate GENFIRE’s ability to outperform conventional tomography algorithms and to further improve the reconstruction quality by refining the experimentally intended tomographic angles. As this method continues to advance with brighter coherent light sources and more efficient data handling, we expect correlative 3D x-ray fluorescence and ptychographic tomography to be a powerful tool for probing a wide range of frozen-hydrated biological specimens, ranging from small prokaryotes such as bacteria, algae, and parasites to large eukaryotes such as mammalian cells, with applications that include understanding cellular responses to environmental stimuli and cell-to-cell interactions.
关键词： GENFIRE,x-ray fluorescence microscopy,frozen-hydrated biological specimens,3D correlative microscopy,x-ray ptychography
更新于2025-09-10 09:29:36