Methods and Applications in Fluorescence最新文献

Absorption and fluorescence spectra of the nitrogen polycyclic aromatic hydrocarbon carbazole (CZL) were analyzed with native cyclodextrins (CD;α,β,γ); derivatizedCD(hydroxypropyl-β-cyclodextrin,HPCD; methyl-β-cyclodextrin,MeCD) and p-sulfonated calixarenes (SCAn, with n = 6 and 8) macrocycles. The results showed a slight increase in the absorbance ofCZLwithCD, but the mixture ofCZLwithSCAshowed lower absorption than the sum of the individual spectra. Also, changes in fluorescence were observed by adding the macrocycles, quenching withSCA, and significant increases withCD. The higher fluorescence enhancement was withHPCDrationalized as a complex formation with 1:1 stoichiometry, with an average value for the association constant (K_A) of (12 ± 1) x 10²M^-1, and a quantum yield ratio between the complexedCZLand freeCZL(Φ^CZL-HPCD/Φ^CZL) of (1.56 ± 0.02) at neutral pH and 25.0 °C. These increases in fluorescence were used as an on-fluorescence switch to develop a supramolecular analytical method forCZLin aqueous samples. The best analytical parameters were inHPCD(LOD = 1.41 ± 0.01 ng mL^-1). The method was validated in aqueous samples of river and tap water with recoveries between 96%-104%. The proposed supramolecular method is quick, direct, selective and represents an alternative and low-cost analysis method.

Here we apply the SUPPOSe algorithm on images acquired using Stimulated Emission Depletion (STED) microscopy with the aim of improving the resolution limit achieved. We processed images of the nuclear pore complex (NPC) from cell lines in which the Nup96 nucleoporin was endogenously labeled. This reference protein forms a ring whose diameter is ∼107 nm with 8 corners ∼42 nm apart from each other. The stereotypic arrangement of proteins in the NPC has been used as reference structures to characterize the performance of a variety of microscopy techniques. STED microscopy images resolve the ring arrangement but not the eightfold symmetry of the NPC. After applying the SUPPOSe algorithm to the STED images, we were able to solve the octagonal structure of the NPC. After processing 562 single NPC, the average radius of the NPC was found to beR= 54.2 ± 2.9 nm, being consistent with the theoretical distances of this structure. To verify that the solutions obtained are compatible with a NPC-type geometry, we rotate the solutions to optimally fit an eightfold-symmetric pattern and we count the number of corners that contain at least one localization. Fitting a probabilistic model to the histogram of the number of bright corners gives an effective labeling efficiency of 31%, which is in agreement with the values reported in for other cell lines and ligands used in Single Molecule Localization microscopy, showing that SUPPOSe can reliably retrieve sub-resolution, nanoscale objects from single acquisitions even in noisy conditions.

We here report the formation of a turbid-gel phase in acrylic cuvettes upon exposure to pure Dimethyl Sulfoxide (DMSO) at room temperature. The observed phenomenon occurred over a 10 h to 14 h incubation in the presence of environmental oxygen. After the turbid gel was removed from the cuvette, it became a white solid exhibiting unique emission behavior. The formation of the turbid-gel phase was accelerated upon exposure to UV 295 LED pulses of light from 6 h to 8 h. Surprisingly, subsequent exposure of the white solid to a few microliters of pure DMSO and vortexing resulted in its transformation into a transparent gel state in just a few minutes, eventually acquiring transparent and liquid properties. Additionally, the white-solid phase can load other molecules, such as Resveratrol and Quercetin, leading to shifts in the respective emission spectra compared with the same molecule in liquid and pure DMSO. These novel findings highlight the interaction between UV photons, oxygen, DMSO and Acrylic, and potentially distort fluorescence spectroscopy experiments.

Forsythoside E is one secondary metabolite ofForsythia suspensa(Thunb.) Vahl. In the study, the interactions between forsythoside E and two types of cholinesterases, acetylcholinesterase and butyrylcholinesterase were investigated in the different conditions. Forsythoside E increased the fluorescence intensity of acetylcholinesterase but quenched the fluorescence of butyrylcholinesterase. Aβ_25-35used in the study may not form complexes with cholinesterases, and did not affect the interaction between forsythoside E and cholinesterases. The charged quaternary group of AsCh interacted with the 'anionic' subsite in acetylcholinesterase, which did not affect the interaction between forsythoside E and acetylcholinesterase. The enhancement rate of forsythoside E to acetylcholinesterase fluorescence from high to low was acid solution (pH 6.4), neutral solution (pH 7.4) and alkaline solution (pH 8.0), while the reduction rate of forsythoside E to butyrylcholinesterase fluorescence was in reverse order. Metal ions may interact with cholinesterases, and increased the effects of forsythoside E to cholinesterases fluorescence, in order that Fe³⁺was the highest, followed by Cu²⁺, and Mg²⁺. A forsythoside E-butyrylcholinesterase complex at stoichiometric ratio of 1:1 was spontaneously formed, and the static quenching was the main quenching mode in the process of forsythoside E binding with butyrylcholinesterase. TheKvalues of two complexes were pretty much the same, suggesting that the interaction between cholinesterases and forsythoside E was almost unaffected by acid-base environment and metal ions. Thennumbers of two cholinesterases approximately equaled to one, indicating that there was only one site on each cholinesterase applicable for forsythoside E to bind to.

Many medical imaging modalities have benefited from recent advances in Machine Learning (ML), specifically in deep learning, such as neural networks. Computers can be trained to investigate and enhance medical imaging methods without using valuable human resources. In recent years, Fluorescence Lifetime Imaging (FLIm) has received increasing attention from the ML community. FLIm goes beyond conventional spectral imaging, providing additional lifetime information, and could lead to optical histopathology supporting real-time diagnostics. However, most current studies do not use the full potential of machine/deep learning models. As a developing image modality, FLIm data are not easily obtainable, which, coupled with an absence of standardisation, is pushing back the research to develop models which could advance automated diagnosis and help promote FLIm. In this paper, we describe recent developments that improve FLIm image quality, specifically time-domain systems, and we summarise sensing, signal-to-noise analysis and the advances in registration and low-level tracking. We review the two main applications of ML for FLIm: lifetime estimation and image analysis through classification and segmentation. We suggest a course of action to improve the quality of ML studies applied to FLIm. Our final goal is to promote FLIm and attract more ML practitioners to explore the potential of lifetime imaging.

Biofluorescence in echinoderms is largely unexplored, and even though the green sea urchinStrongylocentrotus droebachiensisis a well-studied species, the presence and/or function of fluorescence remains very poorly understood. Hyperspectral imaging was conducted on adult sea urchins (N = 380) while fluorospectrometric analysis was conducted on sea urchin coelomic fluid (N = 30). Fluorescence was documented in both the spines and coelomic fluid ofS. droebachiensis. Intact spines exhibited a low intensity green emission (∼550-600 nm), while broken spines averaged a high emission peak in the green spectrum (∼580 nm). Sea urchins produce a red exudate with a pronounced emission peak (∼680 nm) with a shoulder peak (∼730 nm). The sampled coelomic fluid exhibited high variability, with a majority exhibiting a low-level green fluorescence while pronounced emission peaks (N = 5) were found in the red spectrum (∼680 nm). The complex fluorescence produced byS. droebachiensiswarrants further investigation on its applicability for monitoring welfare of sea urchins in aquaculture facilities.

Dopamine (DA) is the most abundant catecholamine neurotransmitter in the brain and plays an extremely essential role in the physiological activities of the living organism. There is a critical need for accurately and efficiently detecting DA levels in organisms in order to reflect physiological states. Carbon nitride quantum dots (C₃N₄) were, in recent years, used enormously as electrochemical and fluorescence probes for the detection of metal ions, biomarkers and other environmental or food impurities due to their unique advantageous optical and electronic properties. 3-Aminophenylboronic acid (3-APBA) can specifically combine with DA through an aggregation effect, providing an effective DA detection method. In this work, 3-APBA modified carbon nitride quantum dots (3-APBA-CNQDs) were synthesized from urea and sodium citrate. The structure, chemical composition and optical properties of 3-APBA-CNQDs were investigated by XRD, TEM, UV-visible, and FT-IR spectroscopy. The addition of DA could induce fluorescence quenching of 3-APBA-CNQDs possibly through the inner filter effect (IFE). 3-APBA-CNQDs shows better selectivity and sensitivity to DA than other interfering substances. By optimizing the experiment conditions, good linearity was obtained at 0.10-51μM DA with a low detection limit of 22.08 nM. More importantly, 3-APBA-CNQDs have been successfully applied for the detection of DA in human urine and blood samples as well as for bioimaging of intracellular DA. This study provides a promising novel method for the rapid detection of DA in real biological samples.

Duvelisib (DUV) is a potent anticancer drug whereas Moxifloxacin (MOX) is an antimicrobial drug with anti-proliferative potency against cancerous cells, which is empirically administered in cancer treatment. DUV and MOX combination is commonly prescribed to combat infections in patients while they are under chemotherapy treatment. This study describes, for the first time, the development of a simple and green synchronous spectrofluorimetric (SSF) method for the simultaneous estimation of DUV and MOX in plasma. DUV and MOX were quantified at 273 and 362 nm, respectively without interference between each other at Δλof 120 nm. The experimental variables influencing fluorescence intensities were thoroughly investigated and the optimum conditions were established. At pH 3.5, the optimum synchronous fluorescence intensity (SFI) was achieved in water solvent by using sodium acetate buffer solution. Calibration curves for DUV and MOX, correlating the SFI with the corresponding drug concentration, were linear in the range of 50-1000 ng mL^-1for both drugs, with good correlation coefficients. The method was extremely sensitive, with limits of detection of 24 and 22 ng mL^-1, and limits of quantitation of 40 and 45 ngmL^-1for DUV and MOX, respectively. The SSF method was validated according to the Food and Drug Administration (FDA) guidelines for validation of analytical procedures, and the validation parameters were acceptable. The proposed SSF method was applied to the pharmacokinetic and bioavailability studies in rats' plasma after single concurrent oral administration of both drugs. The results of the study revealed that caution should be taken with DUV dose when concurrently administered with MOX. The greenness of SSF method was assessed by three different metric tools namely Analytical Eco-scale, Green Analytical Procedure Index, and Analytical Greenness Calculator. The results confirmed that SSF method is an eco-friendly and green analytical approach. In conclusion, the proposed SSF method is a valuable tool for pharmacokinetic/bioavailability studies and therapeutic drug monitoring of simultaneously administered DUV and MOX.

Although the great importance of oral contraceptive agents in birth control, their existence in breast milk became a cause for concern, since infant exposure to these hormones is associated with many health problems. Consequentially, developing a sensitive bioanalytical method for monitoring their concentrations in breast milk is an urgent demand to examine the safety or the risk of these compounds on infants. Levonorgestrel is one of the most common contraceptive hormones under concern. Despite the high sensitivity of the fluorometric methods, detection of Levonorgestrel by them is confined because its structure does not exhibit any fluorescence. For the first time, we proposed a promising click fluorescent probe, 4-azido-7-nitrobenzoxadiazole to react with the alkyne group of Levonorgestrel, to give a highly fluorescent triazole derivative that exhibited strong signal at wavelength of 544 nm after excitation at 470 nm. Reaction parameters impacting the fluorescence were cautiously studied and optimized. The suggested approach has been successfully applied in Levonorgestrel estimation in breast milk samples with linearity of (0.4-80 ng.ml^-1) and low detection limit of 0.12 ng.ml^-1without interferences from any biological components and with mean % recovery of 97.84 ± 2.73. Accuracy, sensitivity, selectivity, simplicity, and low-cost makes this approach a convincing, promising, and appealing alternative over reported analytical methods for Levonorgestrel bioanalysis in different matrices.