Oligonucleotides with sequences of human telomere DNA or thrombin binding aptamer (TBA) are known to form tetraplex structures upon binding the K+ ion. Structural changes associated with the formation of tetraplex assemblies led to the development of potassium-sensing oligonucleotide (PSO) probes, in which two fluorescent dyes were attached to both termini of particular oligonucleotide. The combination of dyes included fluorescence resonance energy transfer (FRET) and excimer emission approaches, and the structural changes upon binding K+ ion could be monitored by a fluorescence technique. These systems showed a very high preference for K+ over Na+ ion, which was suitable for fluorescence imaging of the potassium concentration gradient in a living cell. In the case of human telomere DNA, it was also possible to follow the polymorphism of its tetraplex structures.
Highly efficient cell-free plasma separation from 200 μL of human whole blood was realized via axial migration of blood cells and cross-flow filtration in a microchip. Although various analyses of small volumes of blood have been reported, a large volume of blood is necessary for obtaining blood cells and plasma for the conventional plasma separation technique of centrifugation. A highly efficient plasma separation method using small volumes of blood without hemolysis is an important issue. We developed a plasma separation method based on a microchip with a filter, which utilizes the axial migration of blood cells observed in blood vessels. Clogging and hemolysis on the filter can be prevented by the axial migration of the blood cells. Using this method, 65% of the plasma from 200 μL of whole blood was successfully separated without hemolysis. When the plasma separation microchip interfaced with a micro-ELISA system was applied to C-reactive protein (CRP) analysis, the CRP concentration obtained by the microchip showed good correlation with that obtained by conventional centrifugation. Total analysis time, including plasma separation, was achieved in only 25 min.
In this paper, we describe a method to enhance the fluorescence signal of mutagen detection using SOS response-induced green fluorescence protein (GFP) in genetically modified Escherichia coli using a multi-layered substrate. To generate E. coli that express SOS response-induced GFP, we constructed a plasmid carrying the RecA promoter located upstream of the GFP gene and used it to transform E. coli BL21. The transformed strain was incubated with mitomycin C (MMC), a typical mutagen, and then immobilized on a multi-layered substrate with Ag and a thin Al2O3 layer on a glass slide. Since the multi-layered substrate technique is an optical technique with potential to enhance the fluorescence of fluorophore placed on top of the substrate, the multi-layered substrate was expected to improve the fluorescence signal of mutagen detection. We obtained an average 14-fold fluorescence enhancement of MMC-induced GFP in the concentration range 1 to 1000 ng/ml. In addition, the lower detection limit of MMC was improved using this technique, and was estimated to be 1 ng/ml because of an enlargement of the difference between the blank and the signal of 1 ng/ml of MMC.
In the present work, two aptamer-based probes and related sensor systems were developed with chemiluminescence signaling. The detection was based on “turning-on” chemiluminescence with switching “off” of the resonance energy transfer after the aptamer’s recognition of the target molecule. In this design, a DNA/aptamer duplex linked a chemiluminescence group and a gold nanoparticle together. Only low-intensity chemiluminescence was obtained due to the highly efficient resonance energy transfer. After introducting the target molecule, structure-switching took place with turning off the energy transfer; thus, a restoration and turning on of the chemiluminescence was obtained. The two designs differed in the chemiluminescence groups, since one was a covalently linked luminol molecule, while the other was a conjugated horseradish peroxidase for the catalysis of further chemiluminescence reactions. These schemes provided simple and effective sensing toward a model analyte, adenosine.
We have developed a temperature cycler for polymerase chain reaction (PCR) in a microwell fabricated on a polymer/glass chip. The entire system consisted of three subsystems, which included (1) a thermal conditioner, (2) a proportional-integral-derivative (PID) control signal conditioner and (3) a data acquisition subsystem. The subsystems were regulated coordinately by a ladder logic program written for the programmable logic control (PLC) so that an actual sample temperature could be timed, changed and maintained according to the programmed temperature cycles. The present temperature control system showed high accuracy, stability and minimum overshoot with reduced heating and cooling transition rates. Applicability of the temperature controller to the miniaturized PCR system with reduced volumes of aqueous sample droplets isolated in an oil phase was confirmed by successful amplifications of a target DNA sequence in the microwell.
A novel spectrophotometric flow injection method for determination of silver(I) in a strongly acidic solution containing concentrated copper(II) was developed using a coloring ligand, 4-(3,5-dibromo-2-pyridylazo)-N-ethyl-N-(3-sulfopropyl)aniline (3,5-diBr-PAESA). The method was first investigated by batch method. The interference from copper(II) chelate could be eliminated by the masking effect of EDTA. By utilizing the large formation constant (K = 12.3) of AgBr, one could determine silver(I) as a decrease of absorption by silver(I) chelate due to formation of AgBr by addition of KBr. Based on the results of batch experiments, two types of flow injection analysis (FIA) systems were constructed. Sub-mg dm−3 determination of silver(I) was attained without interference from excess copper(II). The proposed method was successfully applied to determination of silver in a copper plating solution used in a plant to manufacture copper printed circuit boards, where the concentration of silver was critically important in the process control.
A vitamin U-bonded stationary phase was prepared and the retention behavior of inorganic anions was examined using ion chromatography. Inorganic anions were retained on the vitamin U-bonded stationary phase under acidic as well as neutral eluent conditions in the ion-exchange mode. The elution order of the examined anions under neutral eluent conditions was nearly the same as that observed in common ion exchange mode, while the elution order observed under acidic eluent conditions was completely different from that observed in common ion exchange mode. The retention of the analyte anions under the neutral eluent conditions was due to the sulfonium groups of the vitamin U, while protonated primary amino groups caused retention of the analyte anions with different selectivity under acidic conditions. The retention factor of the analyte anions increased with decreasing eluent concentration under both eluent conditions. The present system was applied to the determination of bromide and nitrate contained in seawater.
The ion-pair formation constants (KMLX0/mol−1 dm3) of CdL2+ with Br− or NaL+ with N,N-diethyldithiocarbamate ion (DDTC−) in water were determined potentiometrically at 25°C; ionic strength (I)→0: L denotes 18-crown-6 ether (18C6) and its mono-benzo derivative for the CdBr2-L system and 15-crown-5 ether and 18C6 for the NaDDTC-L one. The formation constant corresponding to the simple salt, NaDDTC, in water was also determined at I→0. Using the log KCdLX0 values of CdLCl+, CdLBr+, CdLPic+, and CdLSO4, then CdL2+ and picrate ion (Pic−) in water have been classified with the hard and soft acids and bases principle, where the values were available in the literature, except for CdLBr+. The same classification was examined in NaX-L systems with X− = DDTC−, trifluoroacetate ion, MnO4−, ReO4−, Pic−, and BPh4− and the AgPic-L one. Consequently, CdL2+, NaL+, and AgL+ were classified as the hard acids, while Pic− and BPh4− as the hard bases. These results reflected the reactivities of the complex ions in ion-pair formation with X− and SO42− in water.
A chemical dosimetry system based on the radiochemical formation of phenol from aqueous benzene solutions was applied to measure the intensity of γ-ray irradiation. Using a simple and sensitive isocratic fluorometric HPLC system, radiochemically generated phenol was determined. The radiochemical formation of phenol was linear up to 100 Gy and the lower limit of detection calculated from the detection limits of phenol was estimated to be 7 mGy. The phenol formation rates were not affected by the oxygen saturation. The chemical dosimetry system investigated in this study was sensitive and was easier to use than traditional chemical dosimeters.
A standard material containing chrysotile asbestos for the validation of x-ray diffractometric quantitation was developed using an asbestos-containing building material i.e., perlite board. The board as the base material was crushed, pulverized, and homogenized. The homogeneity of the powder of perlite board was estimated by analysis of variance. The diffraction intensity values of the crystalline phases and the concentrations of elements determined by x-ray diffractometry and x-ray fluorescence analysis were used for analysis of variance. There is no significant difference between the within-bottle variance and the between-bottle variance, indicating that the powdered perlite board was sufficiently homogenous. The concentration of chrysotile in the material was determined using two methods: an internal standard/x-ray diffractometry method and the x-ray diffractometry/Rietveld refinement. The concentration of chrysotile in the material was determined by an internal standard/x-ray diffractometry method and the material had a chrysotile concentration of 24.1 ± 0.2 mass%.
A promising electrochemical sensor was fabricated by the self-assembling of Pt nanoparticles (nano-Pts) on a chitosan (CS) modified glassy carbon electrode (GCE). A field-emission scanning electron microscope (FE-SEM), transmission electron microscopy (TEM) and electrochemical techniques were used for characterization of these composites. It has been found that nano-Pts are inserted into the CS layer uniformly, and have a larger surface area compared to the chitosan modified glassy carbon electrode. Electrocatalytic experiments for the oxidation of nitrite and the reduction of iodate have shown that nano-Pts/CS/GCE can decrease the over-potential and increase the faradic current, which can be used for the sensitive determination of nitrite and iodate. Moreover, the prepared modified electrode exhibits good reproducibility and stability, and it is possible that this novel electrochemical sensor can be applied in the sensing and/or biosensing field.
A new method for the detection of point mutation in DNA based on the monobase-coded cadmium tellurium nanoprobes and the quartz crystal microbalance (QCM) technique was reported. A point mutation (single-base, adenine, thymine, cytosine, and guanine, namely, A, T, C and G, mutation in DNA strand, respectively) DNA QCM sensor was fabricated by immobilizing single-base mutation DNA modified magnetic beads onto the electrode surface with an external magnetic field near the electrode. The DNA-modified magnetic beads were obtained from the biotin-avidin affinity reaction of biotinylated DNA and streptavidin-functionalized core/shell Fe3O4/Au magnetic nanoparticles, followed by a DNA hybridization reaction. Single-base coded CdTe nanoprobes (A-CdTe, T-CdTe, C-CdTe and G-CdTe, respectively) were used as the detection probes. The mutation site in DNA was distinguished by detecting the decreases of the resonance frequency of the piezoelectric quartz crystal when the coded nanoprobe was added to the test system. This proposed detection strategy for point mutation in DNA is proved to be sensitive, simple, repeatable and low-cost, consequently, it has a great potential for single nucleotide polymorphism (SNP) detection.
Integrating the advantages of screen printing technology with the encouraging electroanalytical characteristic of metallic bismuth, we developed an ultrasensitive and disposable screen-printed bismuth electrode (SPBE) modified with multi-walled carbon nanotubes (MWCNTs) for electrochemical stripping measurements. Metallic bismuth powders and MWCNTs were homogeneously mixed with graphite-carbon ink to mass-prepare screen-printed bismuth electrode doped with multi-walled carbon nanotubes (SPBE/MWCNT). The electroanalytical performance of the prepared SPBE/MWCNT was intensively evaluated by measuring trace Hg(II) with square-wave anodic stripping voltammetry (SWASV). The results indicated that the SPBE modified with 2 wt% MWCNTs could offer a more sensitive response to trace Hg(II) than the bare SPBE. The stripping current obtained at SPBE/MWCNT was linear with Hg(II) concentration in the range from 0.2 to 40 μg/L (R2 = 0.9976), with a detection limit of 0.09 μg/L (S/N = 3) under 180 s accumulation. The proposed “mercury-free” electrode, with extremely simple preparation and ultrahigh sensitivity, holds wide application prospects in both environmental and industrial monitoring.
The effects of light and methyl jasmonate (MJ) on the transcription of biosynthetic genes as well as the accumulation of vindoline and catharanthine in Catharanthus roseus C20hi cell suspensions were studied. t16h (the gene encoding tabersonine 16-hydroxylase) could be induced by light and MJ, whereas d4h (the gene encoding deacetoxyvindoline 4-hydroxylase) could only be induced by light. Quantification by UPLC-MS showed that light significantly increased vindoline production in C20hi cells by about 0.49 – 5.51-fold more than that in controls, with the highest yield being 75.3 ng/g of dry weight. The biosynthesis of vindoline was further enhanced by combining MJ with light. The accumulation of catharanthine was not improved by either light or MJ elicitation. These results suggested that light and MJ could promote vindoline, but not catharanthine accumulation in C20hi cells.
Benzyldimethyltetradecylammonium (BA14+) salts with anionic ligands (X−), such as bis(2-ethylhexyl)sulfosuccinate, bis(2-ethylhexyl)phosphate (BEHP−), and benzotriazole (BTA−) anions, were prepared. These salts were soluble in various organic solvents. The luminescence emission spectra of organic solutions of a red luminescent, tris(1,1,1,2,2,3,3-heptafluoro-7,7-dimethyl-4,6-octadionato)europium(III) complex in the presence of the BA14X’s were recorded. The emission intensity of the Eu(III) complex was increased remarkably by the addition of BA14X (X− = BEHP− and BTA−). This effect can be attributed to the formation of 1:1 X−-adducts of the Eu(III) complex, in which the asymmetry of the ligand field is increased so as to enhance the emission efficiency of the 5D0→5F2 transition. The enhancement effect by BA14X was higher than that of charge-neutral ligands, such as tri-n-octylphosphine oxide and 1,10-phenanthroline, which have been used as second ligands to enhance the emission efficiency of tris(β-diketonato)europium(III) complexes.
The release of perylene from octadecylsilyl (ODS)-silica gel into water using a nonionic surfactant was kinetically studied by single microparticle injection and absorption microspectroscopy techniques. The release of perylene from the porous microparticles significantly depended on the surfactant concentration. The release rate constant was inversely proportional to the microparticle radius; the rate-determining step was the process at the spherical microparticle surface. The mechanism is discussed in terms of the solubilization of perylene at the microparticle surface by the micelle.
Retention behaviors of pyrazine and alkylpyrazines on various stationary phases in reversed-phase liquid chromatography were examined. An abnormal temperature effect on the retention of alkylpyrazines with a mobile phase consisting of acetonitrile and water was observed when changing the column temperature. On the other hand, no similar trend was found with a methanol–water mobile phase. For all the columns investigated in this work, the above tendency to the temperature-dependence was consistently observed, suggesting that the abnormal temperature effect on the retention of alkylpyrazines could be mainly induced by an acetonitrile-based mobile phase.