Methods and Applications in Fluorescence最新文献

Accurate and efficient autofocusing is essential for the automation of fluorescence microscopy, but background noise and shallow depth of field at high magnifications make autofocusing particularly challenging. Here, we present a fast and accurate autofocus algorithm to address these challenges. It is highly effective for high-magnification imaging, while performing equally well for low-magnification imaging tasks. The method is based on the mountain climbing search algorithm and yields improvements on autofocusing precision of up to 200-fold over current methods, whilst offering competitive speed and greatly extended search ranges. Our approach is broadly applicable: it demonstrated good stability and reproducibility across magnifications ranging from 20X to 100X, excels in both live cell imaging and high-resolution fixed sample imaging, and it is compatible with various microscopy techniques without the need for fiducial markers or hardware modifications on existing microscopes. To maximise its accessibility, we constructed a user-friendly interface compatible with the widely used Micromanager software. It generalises well across various imaging modalities and hardware platforms, making it particularly suitable for use in high-resolution screening of candidate drugs.

Ultraviolet (UV) microscopy is a powerful imaging modality that harnesses the shorter wavelengths of UV light to achieve high-resolution imaging and probe molecular-level chemical and structural properties of biological and biomedical specimens, often without the need for extrinsic labelling. Innovations in technologies such as low-cost illuminators, detectors, and new ways of preparing specimens for imaging have led to a better understanding of complex biological systems. Here we review the latest advances and trends in UV microscopy for applications in the life sciences, including histology, cell biology and haemotology. By examining these developments, we highlight the evolving potential of UV and we conclude by considering the future of this longstanding technique.

Multifocal Structured Illumination Microscopy (MSIM) was initially introduced as a parallel version of image scanning microscopy, aiming to enhance the temporal resolution of the imaging process. Beyond its capacity in super-resolution imaging, MSIM exhibits optical sectioning capabilities akin to confocal microscopy, making it well-suited for imaging thick tissue samples. Traditional MSIM reconstruction algorithms rely on digital pinholes to eliminate out-of-focus signals, demanding precise illumination information. However, controlling and accurately reconstructing illumination patterns can be challenging or impractical in certain experimental settings. To address this, our paper proposes a blind reconstruction method for MSIM that circumvents the need for exact illumination information. Leveraging the stability of the standard deviation for each pixel in illumination, this method achieves optical sectioning effectively and provides approximately 1.76 times better resolution than widefield imaging. The efficacy of our proposed blind reconstruction method for both super-resolution imaging and optical sectioning is validated through both simulations and experimental results.

This study explores the fluorescence enhancement of quantum dots (QDs) by gold film over nanospheres (AuFoN) plasmonic substrates, focusing on how a polymer matrix and plasmon resonances of the substrate affect the fluorescence properties of QDs. It was observed that polyvinylpyrrolidone (PVP) facilitated the uniform distribution of QDs on the surface of the AuFoN by simple drop-coating, avoiding aggregation during solvent evaporation. Progressive fluorescence redshifts and intensity enhancement were observed when moving from QDs on glass substrates to planar Au, and most pronouncedly, to nanostructured AuFoN substrates. The fluorescence enhancement was further analyzed by varying the diameter of the polystyrene spheres used in AuFoN fabrication, revealing that substrates based on 600-700 nm spheres provided the strongest fluorescence amplification due to stronger localized electromagnetic fields. Time-resolved fluorescence measurements revealed two primary fluorescence lifetime components for QDs on AuFoN: a short component linked to non-radiative plasmonic energy transfer and a long component representing intrinsic QDs emission. By optimizing sphere size, Au nanostructured films can be tailored to control QDs fluorescence lifetimes and intensity, advancing their use in biosensing, photonics, and other fluorescence-based technologies. This work enhances our understanding of how substrate design and matrix effects impact QDs fluorescence, providing a pathway for precisely engineered Surface Enhanced Fluorescence (SEF) platforms suited to various applications in optical sensing and more general photonics.

Carbon nanodots (CNDs) were synthesized from hazelnut shells using the pyrolysis method in an oven at temperatures ranging from 250 to 400 °C and reaction times between 1 and 3 h. The structural and optical properties of the CNDs, which exhibited strong blue fluorescence under a UV lamp (365 nm), were characterized using UV-vis absorption spectroscopy, fluorescence spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, x-ray diffraction (XRD), x-ray photoelectron spectroscopy (XPS), and transmission electron microscopy (TEM). The quantum yield of the CNDs was determined to be between 2.2% and 7.8%. The effect of various cations on the fluorescence spectra of the CNDs was investigated using fluorescence spectrometry. Among the synthesized CNDs, those prepared via pyrolysis for 3 h at 300 °C, 315 °C, and 350 °C (designated as HS300-3, HS315-3, and HS350-3, respectively) exhibited selective fluorescence quenching in the presence of Fe³⁺, Sn²⁺, and Pd²⁺. These CNDs were applied for the determination of Fe³⁺in tap water and soil samples, while HS315-3 was also employed for the detection of Sn²⁺and Pd²⁺in tap water. The limits of detection (LOD) were 4.5 μg l^-1for Fe³⁺and 15.8 μg l^-1for both Sn²⁺and Pd²⁺. The accuracy of the methods was validated through spiked recovery experiments with tap water samples and by analyzing a certified reference material (CRM-SA-C Sandy Soil C).

Chromium (III), as a significant environmental pollutant, poses a serious threat to human health when accumulated in excess, making it imperative to develop highly sensitive and rapid on-site detection methods. In this study, a Eu³⁺-based metal-organic framework (MOF) was successfully synthesized using a dual-ligand strategy, enabling ratiometric fluorescence detection of Cr(III). During the detection process, distinct changes in the fluorescence ratio were observed, accompanied by a notable color shift from red to blue, providing a clear visual cue. Aromatic π-conjugated organic ligands were selected to enhance the photoluminescence properties, while a second ligand was introduced to optimize the crystal size and pore structure of the MOF. The experimental results demonstrated that the MOF material exhibited outstanding ratiometric fluorescence properties for Cr(III) detection, with high stability. Additionally, the material showed excellent selectivity, anti-interference ability, and sensitivity for detecting Cr(III) in water environments, with a detection range of 0-24 μM and detection limit as low as 59 nM. Further investigation revealed that the changes in ratiometric fluorescence signals were induced by Cr(III)-specific partial collapse of the crystal structure and ligand release. Finally, a MOF-based test strip was developed, where the vivid fluorescence color changes enabled visual detection by the naked eye. The test strip also demonstrated good recovery efficiency in real water samples, further confirming the material's potential as a real-time smart sensor for Cr(III) detection.

The paper considers the temperature sensitivity of luminescence of three polymer types films: polymethyl methacrylate, polystyrene and polylactide doped with core-shell CdSe/CdS/ZnS and Cd_0.9Mn_0.1S/ZnS quantum dots. Films with uniform distribution of quantum dots in the polymer matrix were obtained by the spin-coating method. The influence of the quantum dots and polymer type, as well as their mutual content in the composite, on the thermal sensitivity and thermal stability of the films is considered. The thermal stability of the obtained composites was studied during multiple heating-cooling cycles to 100 and 175 °C, and the conditions for obtaining reusable reversible luminescent thermal sensors were established.

Molybdenum disulfide quantum dots (MoS₂QDs) is a new type of graphite like nanomaterial, which exhibited well chemical stability, unique fluorescence characteristics, and excellent biocompatibility. The conventional hydrothermal synthesis of MoS₂generally requires a long-term reaction at high temperature and high pressure. Herein, we have developed a simple and fast MoS₂QDs synthesis scheme using microwave heating, and further modified the surface of MoS₂QDs using 3-aminophenylboronic acid. The 3- aminophenylboronic acid modified MoS₂QDs (B-MoS₂QDs) were further coated by a zinc-based metal-organic backbone (ZIF-8) in a solution containing zinc ions and 2-methylimidazolium. The constructed nanohybrid B-MoS₂@ZIF-8 were successfully applied to the visualization and rapid detection of bilirubin based on the ratiometric fluorescence changes. The linear range for bilirubin detection is 0.2-75 μmol·l^-1, and detection limit is 0.017 μmol·l^-1.

Cell viability assessment plays a crucial role in biological research, pharmaceutical development, and toxicological identification. Here, we used GelRed, a sensitive and safer nucleic acid dye, to selectively label dead cells with red fluorescence (FL) thus distinguishing dead cells from live ones. Further more, the combined use of GelRed and SYTO 9 (another nucleic acid dye) enabled the clear differentiation in FL spectra between the two physiological statuses. The GelRed and SYTO 9 concentrations were optimized to obtain the highest FL ratio of dead to live cells. The GelRed/SYTO 9-based double staining could quantify the cell viability through flow cytometry analysis, with a good correlation between the detected and theoretical dead cell ratios. Compared with traditional prodium iodide (PI) staining, the GelRed/SYTO 9-based double staining showed high accuracy in quantifying dead cell of low levels. The as-developed staining method could be used in biomedical research to accurately measure the cytotoxic effect of various substances in living cells.

Molybdenum disulfide quantum dots (MoS2 QDs) is a new type of graphite like nanomaterial, which exhibited well chemical stability, unique fluorescence characteristics, and excellent biocompatibility. The conventional hydrothermal synthesis of MoS2 generally requires a long-term reaction at high temperature and high pressure. Herein, we have developed a simple and fast MoS2 QDs synthesis scheme using microwave heating, and further modified the surface of MoS2 QDs using 3-aminophenylboronic acid. The 3- aminophenylboronic acid modified MoS2 QDs (B-MoS2 QDs) were further coated by a zinc-based metal-organic backbone (ZIF-8) in a solution containing zinc ions and 2-methylimidazolium. The constructed nanohybrid B-MoS2@ZIF-8 were successfully applied to the visualization and rapid detection of bilirubin based on the ratiometric fluorescence changes. The linear range for bilirubin detection is 0.2-75 μmol•L-1, and detection limit is 0.017 μmol•L-1.