A gel imaging method was employed to quantitate the GFP that had been subjected to denaturation and degradation treatments. This method is able to differentiate the nativity of GFP by relating the observed changes in the position of fluorescent bands which is unable to be detected using the spectrofluorometric method.
In this paper, ambient total suspended particulates (TSP) with a focus on humic-like substances (HULIS) are characterized based on intensive ground-based field samplings collected in Malaysia during non-haze and haze periods caused by peatland fires on the Indonesian island of Sumatra. Furthermore, concentrations of water-soluble organic carbon (WSOC) and carbon content of HULIS (HULIS-C) were determined, and fluorescence spectra of the HULIS samples were recorded by excitation emission matrix (EEM) fluorescence spectroscopy. The concentrations of WSOC and HULIS-C over the entire period ranged from 4.1 to 24 and 1.3 to 18 μgC m-3, respectively. The concentrations of WSOC and HULIS-C during the peatland fire-induced strong haze periods were over 4.3 and 6.1 times higher, respectively, than the average values recorded during the non-haze periods. Even during the light haze periods, the concentrations of WSOC and HULIS-C were significantly higher than their averages during the non-haze periods. These results indicate that peatland fires induce high concentrations of WSOC, particularly HULIS-C, in ambient TSP at receptor sites. EEM fluorescence spectra identified fulvic-like fluorophores at the highest intensity level in the EEM fluorescence spectra of the haze samples. A peak at excitation/emission (Ex/Em) ≈ (290-330)/(375-425) nm is also observed at high intensity, though this peak is normally associated with marine humic-like fluorophores. It is shown that a peak at Ex/Em ≈ (290-330)/(375-425) nm is not derived from marine sources only; furthermore, peatland fires are shown to be important contributors to HULIS around this peak.
The reactions of 2-chloropyrimidine with methylamine, ethylamine and piperidine gave the corresponding 2-N-methylamino-, 2-N-ethylamino- and 2N- piperidinopyrimidines, respectively. The fluorescence properties of these alkylamino derivatives in chloroform, ethyl acetate, carbon tetrachloride, acetone, ether, ethanol and methanol were studied. All the alkylamino derivatives showed the highest fluorescence intensity in polar protic solvents; thus 2-N-methylaminopyrimidine (highest fluorescence intensity at 377 nm when excited at 282 nm) and 2-N-ethylaminopyrimidine (highest fluorescence intensity at 375 nm, when excited at 286 nm) showed the highest fluorescence in methanol. In ethanol, 2-N-piperidinopyrimidine showed a fluorescence peak at 403 nm when excited at 360 nm and in chloroform it fluoresced at 392 nm when excited at 356 nm.
3-Nitro-2-phenoxypyridine and 3-nitro-2-(4-methyl)phenoxypyridine were obtained when 2-chloro-3-nitropyridine was treated with phenol and p-cresol, respectively. Fluorescence studies were carried out in various solvents, in capped and uncapped conditions and for differing concentrations. Both 3-nitro-2-phenoxypyridine and 3-nitro-2-(4-methyl)phenoxypyridine were fluorescent compounds but 3-Nitro-2-(4-methyl)phenoxypyridine was more fluorescent than 3-nitro-2-phenoxypyridine in all the solvents used. The fluorescence intensity decreased with concentration and time.
In this investigation, the structure of bidentate N,N-Schiff base ligand of vanillin, (E)-4-(((2-amino-5-nitrophenyl)imino)methyl)-2-methoxyphenol (HL) was determined by single crystal X-ray diffraction. The interaction of new [CuL2], [NiL2] and [VOL2] complexes with DNA and BSA was explored through UV-Vis and fluorescence spectroscopy. The electronic spectra changes displayed an isosbestic point for the complexes upon titration with DNA. The Kb values for the complexes [CuL2], [NiL2] and [VOL2] were 2.4×105, 1.9×105 and 4.2×104, respectively. [CuL2] complex was bound more toughly than [NiL2] and [VOL2] complexes. These complexes had a significant interaction with Bovine Serum Albumin (BSA) and the results demonstrated that the quenching mechanism was a static procedure. Also, the complexes interacted with BSA by more than one binding site (n>1). Finally, the theoretical studies were performed using the docking method to calculate the binding constants and recognize the binding site of the DNA and BSA with the complexes. The ligand and complexes including Ni2+, Cu2+ and VO2+ ions were colonized by fungal growth.
One of the non-destructive methods used for the identification and verification of metals is by the energy-dispersive X-ray fluorescence (EDXRF) technique. EDXRF analysis provides several important advantages such as simultaneous determination of the elements present, enable to analyse a very wide concentration range, fast analysis with no tedious sample preparation. The paper shows how this technique is developed and applied in the identification and verification of different grades of stainless steels. Comparison of the results obtained from this analysis with certified reference standards show very small differences between them.
The past decade has seen an increase in the number of research papers on ligand binding to proteins based on fluorescence spectroscopy. In most cases, determination of the binding affinity is made by analyzing the quenching of protein fluorescence induced by the ligand. However, many such articles, even those published in reputed journals, suffer from several mistakes with regard to analysis of fluorescence quenching data. Using the binding of phenylbutazone to human serum albumin as a model, we consider some of these mistakes and show how they affect the values of the association constant. In particular, the failure to correct for the inner filter effect and the use of unsuitable equations are discussed. Ligand binding data presented in these articles should be treated with caution, especially in the absence of data from complementary techniques.
Chlorophyll a fluorescence is increasingly being used as a rapid, non-invasive, sensitive and convenient indicator of photosynthetic performance in marine autotrophs. This review presents the methodology, applications and limitations of chlorophyll fluorescence in marine studies. The various chlorophyll fluorescence tools such as Pulse-Amplitude-Modulated (PAM) and Fast Repetition Rate (FRR) fluorometry used in marine scientific studies are discussed. Various commonly employed chlorophyll fluorescence parameters are elaborated. The application of chlorophyll fluorescence in measuring natural variations, stress, stress tolerance and acclimation/adaptation to changing environment in primary producers such as microalgae, macroalgae, seagrasses and mangroves, and marine symbiotic invertebrates, namely symbiotic sponges, hard corals and sea anemones, kleptoplastic sea slugs and giant clams is critically assessed. Stressors include environmental, biological, physical and chemical ones. The strengths, limitations and future perspectives of the use of chlorophyll fluorescence technique as an assessment tool in symbiotic marine organisms and seaplants are discussed.
The past several decades have seen increasing concern regarding the wide distribution of polycyclic aromatic hydrocarbons (PAHs) in environmental matrices. Primary toxicological data show PAHs' persistent characteristics and possible toxicity effects. Because of this pressing global issue, electroanalytical methods have been introduced. These methods are effective for PAH determination in environmental waters, even outclassing sophisticated analytical techniques such as chromatography, conventional spectrophotometry, fluorescence, and capillary electrophoresis. Herein, the literature published on PAHs is reviewed and discussed with special regard to PAH occurrence. Moreover, the recent developments in electrochemical sensors for PAH determination and the challenges and future outlooks in this field, are also presented.
Chromosome identification is essential for linking sequence and chromosomal maps, verifying sequence assemblies, showing structural variations and tracking inheritance or recombination of chromosomes and chromosomal segments during evolution and breeding programs. Unfortunately, identification of individual chromosomes and chromosome arms has been a major challenge for some economically important crop species with a near-continuous chromosome size range and similar morphology. Here, we developed oligonucleotide-based chromosome-specific probes that enabled us to establish a reference chromosome identification system for oil palm (Elaeis guineensis Jacq., 2n = 32). Massive oligonucleotide sequence pools were anchored to individual chromosome arms using dual and triple fluorescent in situ hybridization (EgOligoFISH). Three fluorescently tagged probe libraries were developed to contain, in total 52,506 gene-rich single-copy 47-mer oligonucleotides spanning each 0.2-0.5 Mb across strategically placed chromosome regions. They generated 19 distinct FISH signals and together with rDNA probes enabled identification of all 32 E. guineensis chromosome arms. The probes were able to identify individual homoeologous chromosome regions in the related Arecaceae palm species: American oil palm (Elaeis oleifera), date palm (Phoenix dactylifera) and coconut (Cocos nucifera) showing the comparative organization and concerted evolution of genomes in the Arecaceae. The oligonucleotide probes developed here provide a valuable approach to chromosome arm identification and allow tracking chromosome transfer in hybridization and breeding programs in oil palm, as well as comparative studies within Arecaceae.
Matched MeSH terms: In Situ Hybridization, Fluorescence
Evidence in crime scenes available in the form of biological stains which cannot be visualized during naked eye examination can be detected by imaging their fluorescence using a combination of excitation lights and suitable filters. These combinations selectively allow the passage of fluorescence light emitted from the targeted stains. However, interference from the fluorescence generated by many of the surface materials bearing the stains often renders it difficult to visualize the stains during forensic photography. This report describes the use of background correction algorithm (BCA) to enhance the visibility of seminal stain, a biological evidence that fluoresces. While earlier reports described the use of narrow band-pass filters for other fluorescing evidences, here, we utilize BCA to enhance images captured using commonly available colour filters, yellow, orange and red. Mean-based contrast adjustment was incorporated into BCA to adjust the background brightness for achieving similarity of images' background appearance, a crucial step for ensuring success while implementing BCA. Experiment results demonstrated the effectiveness of our proposed colour filters' approach using the improved BCA in enhancing the visibility of seminal stains in varying dilutions on selected surfaces.
Fluorescence microscopy is a fundamental technique for the life sciences, where biocompatible and photostable photoluminescence probes in combination with fast and sensitive imaging systems are continually transforming this field. A wide-field time-gated photoluminescence microscopy system customised for ultrasensitive imaging of unique nanoruby probes with long photoluminescence lifetime is described. The detection sensitivity derived from the long photoluminescence lifetime of the nanoruby makes it possible to discriminate signals from unwanted autofluorescence background and laser backscatter by employing a time-gated image acquisition mode. This mode enabled several-fold improvement of the photoluminescence imaging contrast of discrete nanorubies dispersed on a coverslip. It enabled recovery of the photoluminescence signal emanating from discrete nanorubies when covered by a layer of an organic fluorescent dye, which were otherwise invisible without the use of spectral filtering approaches. Time-gated imaging also facilitated high sensitivity detection of nanorubies in a biological environment of cultured cells. Finally, we monitor the binding kinetics of nanorubies to a functionalised substrate, which exemplified a real-time assay in biological fluids. 3D-pseudo colour images of nanorubies immersed in a highly fluorescent dye solution. Nanoruby photoluminescence is subdued by that of the dye in continuous excitation/imaging (left), however it can be recovered by time-gated imaging (right). At the bottom is schematic diagram of nanoruby assay in a biological fluid.
Anoectochilus sp. and Ludisia discolor are known as Jewel orchids. Both species are terrestrial wild orchids that grow in shaded areas of forests. The Jewel orchids are renowned for the beauty of their leaves, which are dark-green laced with silvery or golden veins. The orchids are used as a cure in various parts of Asia. Overharvesting and anthropogenic disturbances threaten the existence of the Jewel orchids in the wild, necessitating human intervention in their survival. An understanding of the structure and adaptations of a plant may assist in its survival when propagated outside of its habitat. In this study, ex vitro leaves of Anoectochilus sp. and L. discolor were subjected to freehand sectioning, and then inspected through brightfield and fluorescence microscopy. The study indicated that all parts of both plants presented typical monocotyledonous characteristics except the leaves. The leaves displayed dorsiventrality with distinct palisade and spongy mesophyll layers. The spongy mesophyll layer contained cells which fluoresced a bright red when exposed to ultraviolet, blue, and green light wavelengths, hinting at the presence of anthocyanins for photoprotection. Cyanidin was detected in the leaves of L. discolor, as enumerated through high performance liquid chromatography (HPLC). The observations indicated that Anoectochilus sp. and L. discolor are well-adapted to live under shaded conditions with minimal exposure to light.
A label -free DNAzyme amplified biosensor is found to be highly selective and sensitive towards fluorescent detection of Pb2+ ions in aqueous media. The DNAzyme complex has designed by the hybridization of the enzyme and substrate strand. In the presence of Pb2+, the DNAzyme activated and cleaved the substrate strand of RNA site (rA) into two oligonucleotide fragments. Further, the free fragment was hybridized with a complementary strand on the surface of MBs. After magnetic separation, SYBER Green I was added and readily intercalate with the dsDNA to gives a bright fluorescence signal with intensity directly proportional to the concentration of Pb2+ions. A detection limit of 5 nM in Pb2+ the detection range 0 to 500 nM was obtained. This label- free fluorescent biosensor has been successfully applied to the determination of environmental water samples. Then results open up the possibility for real-time quantitative detection of Pb2+ with convenient potential applications in the biological and environmental field. Graphical Abstract.
We report the case of a 14-year-old girl who presented with a one-month history of back pain and bilateral lower limb weakness preceded by constitutional symptoms. She neither had a family history of malignancy nor a previous history of trauma. A series of imaging procedures revealed an aggressive lesion of the T12 vertebra with a large soft-tissue component and intraspinal extension leading to spinal cord compression causing cord edema. She underwent urgent posterior instrumentation and fixation of T9 to T12 vertebrae due to worsening neurological deficits. Adjuvant and neoadjuvant chemotherapy with palliative spinal stabilisation were also performed. Features of the lesion were highly consistent with ES on immunohistochemical study and fluorescence in situ hybridization (FISH) analysis for the EWSR1 gene. Postoperatively, both of her lower limbs improved in power and she benefited from regular physiotherapy.
Matched MeSH terms: Fluorescence; In Situ Hybridization, Fluorescence
This paper describes the synthesis of poly(1-aminonaphthalene) and its application as a chemosensor for detection of Fe3+ using the naked eye and a fluorimetric method. The conjugated polymer was synthesized by chemical oxidative polymerization using FeCl3 as a catalyst. The response of the polymer towards various metal ions was investigated using colorimetric detection, and ultraviolet-visible and fluorescence spectroscopies. The polymer displayed high selectivity and sensitivity towards Fe3+ compared with other metal ions. A significant colour change from purple to yellow was observed upon addition of Fe3+ by the naked eye. The polymer also showed a high selectivity and sensitivity 'turn-off' fluorescence response towards Fe3+ ions. A good linear response was obtained for Fe3+ concentrations in the range 10-50 mg L-1 with a detection limit of 1.04 mg L-1 . The proposed chemosensor was applied for determination of Fe3+ content in water samples and satisfactory results were obtained.
Variation at the 3' position of fluorescein via Suzuki-Miyaura cross-coupling with aryl and heteroaryl moieties gave a family of anthofluoresceins whose spectroscopic properties were studied. The 1-methylindole derivative gave the highest quantum yield and was observed to behave as a molecular rotor, displaying marked variations in fluorescent intensities with viscosity and offering possible application in cellular sensing and fluorescent polarisation assays.
In this work, mesoporous TiO2-modified ZnO quantum dots (QDs) were immobilised on a linear low-density polyethylene (LLDPE) polymer using a solution casting method for the photodegradation of tetracycline (TC) antibiotics under fluorescent light irradiation. Various spectroscopic and microscopic techniques were used to investigate the physicochemical properties of the floating hybrid polymer film catalyst (8%-ZT@LLDPE). The highest removal (89.5%) of TC (40 mg/L) was achieved within 90 min at pH 9 due to enhanced water uptake by the LDDPE film and the surface roughness of the hybrid film. The formation of heterojunctions increased the separation of photogenerated electron-hole pairs. The QDs size-dependent quantum confinement effect leads to the displacement of the conduction band potential of ZnO QDs to more negative energy values than TiO2. The displacement generates more reactive species with higher oxidation ability. The highly stable film photocatalyst can be separated easily and can be repeatedly used up to 8 cycles without significant loss in the photocatalytic ability. The scavenging test indicates that the main species responsible for the photodegradation was O2●-. The proposed photodegradation mechanism of TC was demonstrated in further detail based on the intermediates detected by LC-time-of-flight/mass spectrometry (LC/TOF-MS).
Water-driven self-assembly of lipids displays a variety of liquid crystalline phases that are crucial for membrane functions. Herein, we characterize the temperature-induced phase transitions in two compositions of an aqueous self-assembly system of the octyl β-D-glucoside (βGlcOC(8)) system, using steady-state and time-resolved fluorescence measurements. The phase transitions hexagonal ↔ micellar and cubic ↔ lamellar were investigated using tryptophan (Trp) and two of its ester derivatives (Trp-C(4) and Trp-C(8)) to probe the polar headgroup region and pyrene to probe the hydrophobic tail region. The polarity of the headgroup region was estimated to be close to that of simple alcohols (methanol and ethanol) for all phases. The pyrene fluorescence indicates that the pyrene molecules are dispersed among the tails of the hydrophobic region, yet remain in close proximity to the polar head groups. Comparing the present results with our previously reported one for βMaltoOC(12), increasing the tail length of the hexagonal phase from C(8) to C(12) leads to less interaction with pyrene, which is attributed to the more random and wobbling motion of the longer alkyl tail. We measured a reduction (more hydrophobic) in the ratio of the vibronic peak intensities of pyrene (I(1)/I(3)) for the lamellar phase compared to that of the cubic phase. The higher polarity in the cubic phase can be correlated to the nature of its interface, which curves toward the bulk water. This geometry also explains the slight reduction in polarity of the headgroup region compared to the other phases. Upon the addition of Trp-C(8), the fluorescence lifetime of pyrene is reduced by 28% in the lamellar and cubic phases, whereas the I(1)/I(3) value is only slightly reduced. The results reflect the dominant role of dynamic interaction mechanism between the C(8) chain of Trp-C(8) and pyrene. This mechanism may be important for these two phases since they participate in the process of membrane fusion. Both lipid compositions show completely reversible temperature-induced phase transitions, reflecting the thermodynamic equilibrium structures of their mesophases. Probing both regions of the different lipid phases reveals a large degree of heterogeneity and flexibility of the lipid self-assembly. These properties are crucial for carrying out different biological functions such as the ability to accommodate various molecular sizes.
A sequencing batch reactor (SBR) seeded with flocculated sludge and fed with synthetic wastewater was operated for an enhanced biological phosphorus removal (EBPR) process. Eight weeks after reactor startup, sludge granules were observed. The granules had a diameter of 0.5 to 3.0 mm and were brownish in color and spherical or ellipsoidal in shape. No significant change was observed in sludge granule size when operational pH was changed from 7 to 8. The 208-day continuous operation of the SBR showed that sludge granules were stably maintained with a sludge volume index (SVI) between 30 to 55 mL/g while securing a removal efficiency of 83% for carbon and 97% for phosphorus. Fluorescent in situ hybridization (FISH) confirmed the enrichment of polyphosphate accumulating organisms (PAOs) in the SBR. The observations of sludge granulation in this study encourage further studies in the development of granules-based EBPR process.
Matched MeSH terms: In Situ Hybridization, Fluorescence