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Elucidating endotoxin-biomolecule interactions with FRET: extending the frontiers of their supramolecular complexation

Obeng EM, Dullah EC, Razak NSA, Danquah MK, Budiman C, Ongkudon CM

J Biol Methods, 2017;4(2):e71.
PMID: 31453229 DOI: 10.14440/jbm.2017.172

Endotoxin has been one of the topical chemical contaminants of major concern to researchers, especially in the field of bioprocessing. This major concern of researchers stems from the fact that the presence of Gram-negative bacterial endotoxin in intracellular products is unavoidable and requires complex downstream purification steps. For instance, endotoxin interacts with recombinant proteins, peptides, antibodies and aptamers and these interactions have formed the foundation for most biosensors for endotoxin detection. It has become imperative for researchers to engineer reliable means/techniques to detect, separate and remove endotoxin, without compromising the quality and quantity of the end-product. However, the underlying mechanism involved during endotoxin-biomolecule interaction is still a gray area. The use of quantitative molecular microscopy that provides high resolution of biomolecules is highly promising, hence, may lead to the development of improved endotoxin detection strategies in biomolecule preparation. Förster resonance energy transfer (FRET) spectroscopy is one of the emerging most powerful tools compatible with most super-resolution techniques for the analysis of molecular interactions. However, the scope of FRET has not been well-exploited in the analysis of endotoxin-biomolecule interaction. This article reviews endotoxin, its pathophysiological consequences and the interaction with biomolecules. Herein, we outline the common potential ways of using FRET to extend the current understanding of endotoxin-biomolecule interaction with the inference that a detailed understanding of the interaction is a prerequisite for the design of strategies for endotoxin identification and removal from protein milieus.

Matched MeSH terms: Fluorescence Resonance Energy Transfer
Intraoral appliances for in situ oral biofilm growth: a systematic review

Abdullah N, Al-Marzooq F, Mohamad S, Abd Rahman N, Chi Ngo H, Perera Samaranayake L

J Oral Microbiol, 2019;11(1):1647757.
PMID: 31489127 DOI: 10.1080/20002297.2019.1647757

Background: Oral biofilms are the root cause of major oral diseases. As in vitro biofilms are not representative of the intraoral milieu, various devices have been manufactured over the years to develop Appliance Grown Oral Biofilm (AGOB). Objective: To review various intraoral appliances used to develop AGOB for microbiological analysis, and to judge the optimal means for such analyses. Design: Four databases (PubMed, Science Direct, Scopus and Medline) were searched by two independent reviewers, and articles featuring the key words 'device' OR 'splint' OR 'appliance'; 'Oral biofilm' OR 'dental plaque'; 'in vivo' OR 'in situ'; 'Microbiology' OR 'Bacteria' OR 'microbiome'; were included. The standard Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) were adopted for data gathering. Results: Of the 517 articles which met the initial inclusion criteria, 24 were deemed eligible for review. The age of the AGOB, sampled at various intervals, ranged from 30 min to 28 days. The most commonly used microbiome analytical methods were fluorescence microscopy, total cell count using conventional, and molecular tools including Next Generation Sequencing (NGS) platforms. Conclusions: No uniformly superior method for collecting AGOB could be discerned. NGS platforms are preferable for AGOB analyses.

Matched MeSH terms: Microscopy, Fluorescence
Fulltext Influence of Sintering Temperature of Kaolin, Slag, and Fly Ash Geopolymers on the Microstructure, Phase Analysis, and Electrical Conductivity

Zulkifli NNI, Abdullah MMAB, Przybył A, Pietrusiewicz P, Salleh MAAM, Aziz IH, et al.

Materials (Basel), 2021 Apr 26;14(9).
PMID: 33925777 DOI: 10.3390/ma14092213

This paper clarified the microstructural element distribution and electrical conductivity changes of kaolin, fly ash, and slag geopolymer at 900 °C. The surface microstructure analysis showed the development in surface densification within the geopolymer when in contact with sintering temperature. It was found that the electrical conductivity was majorly influenced by the existence of the crystalline phase within the geopolymer sample. The highest electrical conductivity (8.3 × 10-4 Ωm-1) was delivered by slag geopolymer due to the crystalline mineral of gehlenite (3Ca2Al2SiO7). Using synchrotron radiation X-ray fluorescence, the high concentration Ca boundaries revealed the appearance of gehlenite crystallisation, which was believed to contribute to development of denser microstructure and electrical conductivity.

Matched MeSH terms: Fluorescence
Fulltext Biofilm formation and granule properties in anaerobic digestion at high salinity

Gagliano MC, Ismail SB, Stams AJM, Plugge CM, Temmink H, Van Lier JB

Water Res, 2017 09 15;121:61-71.
PMID: 28511041 DOI: 10.1016/j.watres.2017.05.016

For the anaerobic biological treatment of saline wastewater, Anaerobic Digestion (AD) is currently a possibility, even though elevated salt concentrations can be a major obstacle. Anaerobic consortia and especially methanogenic archaea are very sensitive to fluctuations in salinity. When working with Upflow Sludge Blanket Reactor (UASB) technology, in which the microorganisms are aggregated and retained in the system as a granular biofilm, high sodium concentration negatively affects aggregation and consequently process performances. In this research, we analysed the structure of the biofilm and granules formed during the anaerobic treatment of high salinity (at 10 and 20 g/L of sodium) synthetic wastewater at lab scale. The acclimated inoculum was able to accomplish high rates of organics removal at all the salinity levels tested. 16S rRNA gene clonal analysis and Fluorescence In Situ Hybridization (FISH) analyses identified the acetoclastic Methanosaeta harundinacea as the key player involved acetate degradation and microbial attachment/granulation. When additional calcium (1 g/L) was added to overcome the negative effect of sodium on microbial aggregation, during the biofilm formation process microbial attachment and acetate degradation decreased. The same result was observed on granules formation: while calcium had a positive effect on granules strength when added to UASB reactors, Methanosaeta filaments were not present and the degradation of the partially acidified substrate was negatively influenced. This research demonstrated the possibility to get granulation at high salinity, bringing to the forefront the importance of a selection towards Methanosaeta cells growing in filamentous form to obtain strong and healthy granules.

Matched MeSH terms: In Situ Hybridization, Fluorescence
Fluorescence "turn-off/turn-on" biosensing of metal ions by gold nanoclusters, folic acid and reduced graphene oxide

Wong XY, Quesada-González D, Manickam S, Muthoosamy K

Anal Chim Acta, 2021 Aug 29;1175:338745.
PMID: 34330444 DOI: 10.1016/j.aca.2021.338745

Metal ions homeostasis plays an important role in biological processes. The ability to detect the concentration of metal ions in biological fluids is often challenged by the obvious interference or competitive binding nature of other alkaline metals ions. Common analytical techniques employed for metal ions detection are electrochemical, fluorescence and colorimetric methods. However, most reported metal ions sensors are complicated, time-consuming and involve costly procedures with limited effectiveness. Herein, a nanobiosensor for detecting sodium and potassium ions using folic acid-functionalised reduced graphene oxide-modified RNase A gold nanoclusters (FA-rGO-RNase A/AuNCs) based on fluorescence "turn-off/turn-on" is presented. Firstly, a facile and optimised protocol for the fabrication of RNase A/AuNCs is developed. The activity of RNase A protein after the formation of RNase A/AuNCs is studied. RNase A/AuNCs is then loaded onto FA-rGO, in which FA-rGO is used as a potential carrier and fluorescence quencher for RNase A/AuNCs. Finally, a fluorescence "turn-on" sensing strategy is developed using the as-synthesised FA-rGO-RNase A/AuNCs to detect sodium and potassium ions. The developed nanobiosensor revealed an excellent sensing performance and meets the sensitivity required to detect both sodium and potassium ions. To the best of our knowledge, this is the first work done on determining the RNase A protein activity in RNase A/AuNCs and exploring the potential application of RNase A/AuNCs as a metal ion sensor. This work serves as a proof-of-concept for combining the potential of drug delivery, active targeting and therapy on cancer cells, as well as biosensing of metal ions into a single platform.

Matched MeSH terms: Spectrometry, Fluorescence
Fulltext Cloning and mutagenetic modification of the firefly luciferase gene and its use for bioluminescence microscopy of engrailed expression during Drosophila metamorphosis

Ogoh K, Akiyoshi R, Suzuki H

Biochem Biophys Rep, 2020 Sep;23:100771.
PMID: 32490216 DOI: 10.1016/j.bbrep.2020.100771

Bioluminescence microscopy is an area attracting considerable interest in the field of cell biology because it offers several advantages over fluorescence microscopy, including no requirement for excitation light and being phototoxicity free. This method requires brighter luciferase for imaging; however, suitable genetic resource material for this purpose is not available at present. To achieve brighter bioluminescence microscopy, we developed a new firefly luciferase. Using the brighter luciferase, a reporter strain of Drosophila Gal4-UAS (Upstream Activating Sequence) system was constructed. This system demonstrated the expression pattern of engrailed, which is a segment polarity gene, during Drosophila metamorphosis by bioluminescence microscopy, and revealed drastic spatiotemporal change in the engrailed expression pattern during head eversion in the early stage of pupation.

Matched MeSH terms: Microscopy, Fluorescence
Determination of pyronaridine in blood plasma by high-performance liquid chromatography for application in clinical pharmacological studies

Jayaraman SD, Ismail S, Nair NK, Navaratnam V

J Chromatogr B Biomed Sci Appl, 1997 Mar 07;690(1-2):253-7.
PMID: 9106050

A method is described for the determination of pyronaridine in plasma using high-performance liquid chromatography with fluorescence detection. The method involves liquid-liquid extraction with phosphate buffer (pH 6.0, 0.05 M) and diethyl ether-hexane (70:30%, v/v) and chromatographic separation on a C18 column (Nucleosil, 250 x 4.6 mm I.D., 5 microns particle size) with acetonitrile-0.05 M phosphate buffer pH 6.0 (60:40%, v/v) as the mobile phase (1 ml/min) and detection by fluorescence (lambda ex = 267 nm, lambda em = 443 nm). The detector response is linear up to 1000 ng and the overall recoveries of pyronaridine and quinine were 90.0 and 60.3%, respectively. The assay procedure was adequately sensitive to measure 10 ng/ml pyronaridine in plasma samples with acceptable precision (< 15% C.V.). The method was found to be suitable for use in clinical pharmacological studies.

Matched MeSH terms: Spectrometry, Fluorescence
Fulltext XRF determination of major elemental contents of clay samples from north-west Peninsular Malaysia

Ahmad Saat, Zaini Hamzah, Zaharidah Abu Bakar

Journal of Nuclear and Related Technologies, 2009;2009(1):230-236.
MyJurnal

Being an imperative material for man either used as building materials, pottery or as components in material industry and technology, knowledge of clays elemental contents is important. In the present study ten clay samples obtained from various locations in North-West Peninsular Malaysia were used. Majority of the clays were economically manufactured to be used as building materials or pottery. The objective of study was to determine the main elemental contents of the samples, and relate the results to the types of minerals, as well as to compare them with clays from other studies. In the study X-ray Fluorescence (XRF) coupled to samples dilution method and standard calibration samples was used. The elements detected in the study were Si, Al, Fe, Ti, K and Ca. Depending on locations, the percentage concentration ranged between 24.8 – 32.4 for Si, 10.8 – 19.0 for Al, 0.09 – 2.12 for Fe, 0.08 – 1.13 for Ti, 0.45 – 3.39 for K and trace amount of Ca and P. However, Mg that normally found in typical clay was not found in the studied samples. Comparing the oxide of the major elements with other studies, it was found that the clay samples contained mixtures of kaolinite (two-layered structure) and illite (three-layered structure).

Matched MeSH terms: Fluorescence
Fulltext Quantitative analysis of Roundup Ready soybean content in soy-derived food and animal feed by using real-time PCR incorporated with cloned DNA fragments

International Food Research Journal, 2011;18(2):507-514.
MyJurnal

Malaysia, Biosafety Bill 2006 was approved by Parliament in July 2007, and labeling legislation will be implemented soon. In this study, duplex polymerase chain reaction (PCR) was carried out to detect
endogenous soybean lectin gene and exogenous cp4-epsps (5’-enolpyruvylshikimate-3-phospate synthase) gene simultaneously. Additionally, real-time PCR utilizing SYBR Green fluorescence dye were established for the quantitative analysis of Roundup Ready soybean (RRS), which is based on the two established calibration curve from cloned fragment of cp4-epsps gene and lectin gene respectively. Approximately, 39.5% (45/114) of the samples examined in this study contain RRS, animal feeds (31), processed food (13) and raw soybean (1). Additionally, 75.6% (34/45) of the positive samples were found contained RRS above 0.9%. The sensitive GMO quantitative approach described in this study enable the analysis of various samples and this will facilitate the labeling process.

Matched MeSH terms: Fluorescence
A Comparative Analysis of Protein Stabilizing Potential of Honey and Simulated Honey Sugar Cocktail

Wong YH, Kadir HA, Tayyab S

Protein Pept Lett, 2016;23(10):898-904.
PMID: 27586182

Urea and thermal denaturations of bovine serum albumin (BSA) were studied in the absence and the presence of honey or simulated honey sugar cocktail (SHSC) using far-UV CD and ANS fluorescence spectroscopy. Presence of 20% (w/v) honey or SHSC in the incubation mixture shifted the urea transition curve towards higher urea concentrations, being higher in the presence of honey and transformed the two-step, three-state transition into a single-step, two-state transition. A comparison of the far-UV CD and the ANS fluorescence spectra of 4.6 M urea-denatured BSA (U-BSA) in the absence and the presence of 20% (w/v) honey or SHSC suggested greater stabilizing potential of honey than SHSC, as U-BSA maintained native like conformation in the presence of 20% (w/v) honey. Furthermore, thermal transition curves of BSA were also shifted towards higher temperature range in the presence of 20% (w/v) SHSC and honey, showing greater shift in the presence of honey. The far-UV CD spectra of the heat-denatured BSA also showed greater stabilization in the presence of honey. Taken together all these results suggested greater protein stabilizing potential of honey than SHSC against chemical and thermal denaturations of BSA.

Matched MeSH terms: Spectrometry, Fluorescence
Pontamine fast scarlet 4B bifluorescence and measurements of cellulose microfibril angles

Thomas J, Idris NA, Collings DA

J Microsc, 2017 10;268(1):13-27.
PMID: 28654160 DOI: 10.1111/jmi.12582

Pontamine fast scarlet 4B is a red paper and textiles dye that has recently been introduced as a fluorescent probe for plant cell walls. Pontamine exhibits bifluorescence, or fluorescence dependent on the polarization of the excitation light: Because cellulose is aligned within the cell wall, pontamine-labelled cell walls exhibit variable fluorescence as the excitation polarization is modulated. Thus, bifluorescence measurements require polarized excitation that can be directly or indirectly modulated. In our confocal microscopy observations of various cellulose samples labelled with pontamine, we modulated excitation polarization either through sample rotation or by the confocal's scanfield rotation function. This variably rotated laser polarizations on Leica confocal microscopes, but not those from other makers. Beginning with samples with directly observable microfibril orientations, such as purified bacterial cellulose, the velamen of orchid roots and the inner S2 layer of radiata pine compression wood, we demonstrate that modelling the variations in pontamine fluorescence with a sine curve can be used to measure the known microfibril angles. We then measured average local microfibril angles in radiata pine samples, and showed similar microfibril angles in compression and normal (opposite) wood. Significantly, bifluorescence measurements might also be used to understand the degree of local cellulose alignment within the cell wall, as opposed to variations in the overall cellulose angle.

Matched MeSH terms: Fluorescence
Fulltext Quantification of intracellular payload release from polymersome nanoparticles

Scarpa E, Bailey JL, Janeczek AA, Stumpf PS, Johnston AH, Oreffo RO, et al.

Sci Rep, 2016 07 11;6:29460.
PMID: 27404770 DOI: 10.1038/srep29460

Polymersome nanoparticles (PMs) are attractive candidates for spatio-temporal controlled delivery of therapeutic agents. Although many studies have addressed cellular uptake of solid nanoparticles, there is very little data available on intracellular release of molecules encapsulated in membranous carriers, such as polymersomes. Here, we addressed this by developing a quantitative assay based on the hydrophilic dye, fluorescein. Fluorescein was encapsulated stably in PMs of mean diameter 85 nm, with minimal leakage after sustained dialysis. No fluorescence was detectable from fluorescein PMs, indicating quenching. Following incubation of L929 cells with fluorescein PMs, there was a gradual increase in intracellular fluorescence, indicating PM disruption and cytosolic release of fluorescein. By combining absorbance measurements with flow cytometry, we quantified the real-time intracellular release of a fluorescein at a single-cell resolution. We found that 173 ± 38 polymersomes released their payload per cell, with significant heterogeneity in uptake, despite controlled synchronisation of cell cycle. This novel method for quantification of the release of compounds from nanoparticles provides fundamental information on cellular uptake of nanoparticle-encapsulated compounds. It also illustrates the stochastic nature of population distribution in homogeneous cell populations, a factor that must be taken into account in clinical use of this technology.

Matched MeSH terms: Fluorescence
Determination of optimum harvest maturity and non-destructive evaluation of pod development and maturity in cacao (Theobroma cacao L.) using a multiparametric fluorescence sensor

Tee YK, Balasundram SK, Ding P, M Hanif AH, Bariah K

J Sci Food Agric, 2019 Mar 15;99(4):1700-1708.
PMID: 30206959 DOI: 10.1002/jsfa.9359

BACKGROUND: A series of fluorescence indices (anthocyanin, flavonol, chlorophyll and nitrogen balance) were deployed to detect the pigments and colourless flavonoids in cacao pods of three commercial cacao (Theobroma cacao L.) genotypes (QH1003, KKM22 and MCBC1) using a fast and non-destructive multiparametric fluorescence sensor. The aim was to determine optimum harvest periods (either 4 or 5 months after pod emergence) of commercial cacao based on fluorescence indices of cacao development and bean quality.
RESULTS: As pod developed, cacao exhibited a rise with the peak of flavonol occurring at months 4 and 5 after pod maturity was initiated while nitrogen balance showed a decreasing trend during maturity. Cacao pods contained high chlorophyll as they developed but chlorophyll content declined significantly on pods that ripened at month 5.
CONCLUSION: Cacao pods harvested at months 4 and 5 can be considered as commercially-ready as the beans have developed good quality and comply with the Malaysian standard on cacao bean specification. Thus, cacao pods can be harvested earlier when they reach maturity at month 4 after pod emergence to avoid germinated beans and over fermentation in ripe pods harvested at month 5. © 2018 Society of Chemical Industry.

Matched MeSH terms: Fluorescence
Fulltext EDX-SEM-XRF data from selected Precambrian Basement Complex rock samples in part of Southwestern Nigeria

Kayode JS, Yusup Y, Nawawi MNM, Ariffin KS, Kalil AE, Tagwa MG

Data Brief, 2018 Oct;20:1525-1531.
PMID: 30258956 DOI: 10.1016/j.dib.2018.09.014

Energy Dispersive X-ray Analysis, EDX mapping, Scanning Electron Microscope, SEM, together with X-ray Fluorescence Analysis, XRF, was carried out to extract the needed data from some metamorphic rock samples in part of the Nigerian Southwestern Precambrian Basement Complex, NSPBC. The foremost aim is to obtain the detail subsurface geological structures of the rocks within the area and to enhanced understanding of the processes and the types of metamorphic evolution in the area. The techniques involved qualitative and quantitative data analysis of the major, minor and radioactive elements present in the samples of rocks analyzed. The data helped to experimentally evaluate the rocks microstructures, and to also explore the development of magmatic and metamorphic mechanisms for the recognition of textual associations in the area. Applications of the EDX, SEM, and XRF data analysis are effortlessly done to determine the varied mixtures of Si, Al, Ca, Fe, K, Mg, and Na, in the presence of O existing in the rocks samples.The data helped in the classification and perceptive of these rocks and it was considered as a necessary tool in the knowledge of the metamorphism and origin of the Basement Complex rocks through measurement of the intensity of the emitted X-ray and its characteristics.

Matched MeSH terms: Fluorescence
Fulltext Hydrogel Fluorescence Microsensor with Fluorescence Recovery for Prolonged Stable Temperature Measurements

Hashim H, Maruyama H, Akita Y, Arai F

Sensors (Basel), 2019 Nov 29;19(23).
PMID: 31795304 DOI: 10.3390/s19235247

This work describes a hydrogel fluorescence microsensor for prolonged stable temperature measurements. Temperature measurement using microsensors has the potential to provide information about cells, tissues, and the culture environment, with optical measurement using a fluorescent dye being a promising microsensing approach. However, it is challenging to achieve stable measurements over prolonged periods with conventional measurement methods based on the fluorescence intensity of fluorescent dye because the excited fluorescent dye molecules are bleached by the exposure to light. The decrease in fluorescence intensity induced by photobleaching causes measurement errors. In this work, a photobleaching compensation method based on the diffusion of fluorescent dye inside a hydrogel microsensor is proposed. The factors that influence compensation in the hydrogel microsensor system are the interval time between measurements, material, concentration of photo initiator, and the composition of the fluorescence microsensor. These factors were evaluated by comparing a polystyrene fluorescence microsensor and a hydrogel fluorescence microsensor, both with diameters of 20 µm. The hydrogel fluorescence microsensor made from 9% poly (ethylene glycol) diacrylate (PEGDA) 575 and 2% photo initiator showed excellent fluorescence intensity stability after exposure (standard deviation of difference from initial fluorescence after 100 measurement repetitions: within 1%). The effect of microsensor size on the stability of the fluorescence intensity was also evaluated. The hydrogel fluorescence microsensors, with sizes greater than the measurement area determined by the axial resolution of the confocal microscope, showed a small decrease in fluorescence intensity, within 3%, after 900 measurement repetitions. The temperature of deionized water in a microchamber was measured for 5400 s using both a thermopile and the hydrogel fluorescence microsensor. The results showed that the maximum error and standard deviation of error between these two sensors were 0.5 °C and 0.3 °C, respectively, confirming the effectiveness of the proposed method.

Matched MeSH terms: Fluorescence
Combination mode of antimalarial drug mefloquine and human serum albumin: Insights from spectroscopic and docking approaches

Musa KA, Ridzwan NFW, Mohamad SB, Tayyab S

Biopolymers, 2020 Feb;111(2):e23337.
PMID: 31691964 DOI: 10.1002/bip.23337

The interaction between mefloquine (MEF), the antimalarial drug, and human serum albumin (HSA), the main carrier protein in blood circulation, was explored using fluorescence, absorption, and circular dichroism spectroscopic techniques. Quenching of HSA fluorescence with MEF was characterized as static quenching and thus confirmed the complex formation between MEF and HSA. Association constant values for MEF-HSA interaction were found to fall within the range of 3.79-5.73 × 104 M-1 at various temperatures (288, 298, and 308 K), which revealed moderate binding affinity. Hydrogen bonds and hydrophobic interactions were predicted to connect MEF and HSA together in the MEF-HSA complex, as deduced from the thermodynamic data (ΔS = +133.52 J mol-1 K-1 and ΔH = +13.09 kJ mol-1 ) of the binding reaction and molecular docking analysis. Three-dimensional fluorescence spectral analysis pointed out alterations in the microenvironment around aromatic amino acid (tryptophan and tyrosine) residues of HSA consequent to the addition of MEF. Circular dichroic spectra of HSA in the wavelength ranges of 200-250 and 250-300 nm hinted smaller changes in the protein's secondary and tertiary structures, respectively, induced by MEF binding. Noncovalent conjugation of MEF to HSA bettered protein thermostability. Site marker competitive drug displacement results suggested HSA Sudlow's site I as the MEF binding site, which was also supported by molecular docking analysis.

Matched MeSH terms: Spectrometry, Fluorescence
Tio2 nanoparticles coated with porphyrin dye thin film as fluorescence gas sensor

Nurul Huda Yusoff, Muhamad Mat Salleh, Muhammad Yahaya

Sains Malaysiana, 2008;37:233-237.

This research explores the possibility of using fluorescence technique to detect the presence of volatile organic compounds based on a single sensing material. The material used was TiO2 nanoparticles coated with porphyrin dye. The TiO2 nanoparticles colloid is in a sol-gel form synthesized from titanium (IV) ethoxide in ethanol with addition of kalium chloride (KCl) as stabilizer. TiO2 nanoparticles were then coated with porphyrin dye, Manganase (III) 5,10,15,20 tetra (4-pyridyl)-21H, 23H porphine chloride tetrakis (metachloride). The coated nanoparticles were deposited on quartz substrate using self-assembly through dip coating technique. The sensing properties of the thin film toward volatile organic compounds; ethanol, acetone, cyclohexane and 2-propanol were studied using luminescence spectrometer. It was found that the thin film produced different emission spectra peaks for different volatile organic compounds (VOCs). Hence, it eases chemical identification process and potentially be use as fluorescence gas sensor.

Matched MeSH terms: Fluorescence
Fulltext Strength Development and Elemental Distribution of Dolomite/Fly Ash Geopolymer Composite under Elevated Temperature

Azimi EA, Abdullah MMAB, Vizureanu P, Salleh MAAM, Sandu AV, Chaiprapa J, et al.

Materials (Basel), 2020 Feb 24;13(4).
PMID: 32102345 DOI: 10.3390/ma13041015

A geopolymer has been reckoned as a rising technology with huge potential for application across the globe. Dolomite refers to a material that can be used raw in producing geopolymers. Nevertheless, dolomite has slow strength development due to its low reactivity as a geopolymer. In this study, dolomite/fly ash (DFA) geopolymer composites were produced with dolomite, fly ash, sodium hydroxide, and liquid sodium silicate. A compression test was carried out on DFA geopolymers to determine the strength of the composite, while a synchrotron Micro-Xray Fluorescence (Micro-XRF) test was performed to assess the elemental distribution in the geopolymer composite. The temperature applied in this study generated promising properties of DFA geopolymers, especially in strength, which displayed increments up to 74.48 MPa as the optimum value. Heat seemed to enhance the strength development of DFA geopolymer composites. The elemental distribution analysis revealed exceptional outcomes for the composites, particularly exposure up to 400 °C, which signified the homogeneity of the DFA composites. Temperatures exceeding 400 °C accelerated the strength development, thus increasing the strength of the DFA composites. This appears to be unique because the strength of ordinary Portland Cement (OPC) and other geopolymers composed of other raw materials is typically either maintained or decreases due to increased heat.

Matched MeSH terms: Fluorescence
DNA-templated silver nanocluster for live-intracellular FOXP3 detection

Lee SY, Fazlina N, Tye GJ

Anal Biochem, 2019 09 15;581:113352.
PMID: 31260647 DOI: 10.1016/j.ab.2019.113352

DNA-templated silver nanocluster (AgNC), a new promising fluorescence probe has gained importance in biosensing and bioimaging in recent years. We employed a label-free AgNC to detect an intracellular transcription factor known as forkhead box p3 (FOXP3), which is the master regulator of regulatory T cells (Tregs) suppressive function. We developed an optimized method for the detection of messenger ribonucleic acid (mRNA) of FOXP3 by hybridizing AgNC and G-rich to the target FOXP3 mRNA of a MCF-7 cells. MCF-7 cells are chosen as a model as it readily expresses FOXP3. The hybridized samples were examined with UV illuminator and further verified with fluorescence spectroscopy, fluorescence microscope and flow cytometry. The successful hybridization of a three-way junction with AgNC, G-rich and mRNA FOXP3 target generated an improved fluorescence intensity with a spectral shift. We have successfully delivered the green fluorescing AgNC and G-rich into MCF-7 cells, producing a shift to red fluorescing cells corroborated by flow cytometry results. In summary, our approach enables the detection of intracellular FOXP3 nucleic acid and holds considerable potential in establishing a non-lethal intracellular detection system which would be crucial for the isolation of regulatory T-cells (Tregs) when combined with other cell surface markers.

Matched MeSH terms: Spectrometry, Fluorescence
Inadvertent expression of dengue 2 virus NS3-EGFP fusion protein in Escherlchla coll using the pEGFP-N 1 mammalian expression vector

Norazizah S, AbuBakar S

JUMMEC, 1999;4:41-46.

Dengue 2 New Guinea C (NGC) virus NS3 protein, a potentially important virulence factor was cloned to the N-terminus of the Aeqirorea victoria enhanced green fluorescent protein (EGFP) using the pEGFP-N1 mammalian expression vector. During amplification of the recombinant plasmid in E. coli, transformants expressing the EGFP were detected in vivo when viewed using fluorescence microscopy. This inadvertent expression of the recombinant fusion protein was confirmed further by detection of the T7.Tag peptide cloned to the aluino terminal of the fusion protein using T 7.Tag specific monoclonal antibody. These findings represent perhaps the first reported expression of the T7.Tag-NS3-EGFP fusion protein using the pEGFP-N1 mammalian expression vector in E. coli. KEYWORDS: Dengue, NS3, pEGFP-N1, fusion protein.

Matched MeSH terms: Fluorescence

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