Fluorimetric determination of nucleic acid using the enhancement of terbium-gadolinium-nucleic acid-cetylpyridine bromide system

It is found that the fluorescence intensity of Tb-cetylpyridine bromide (CPB)-nucleic acid system can be enhanced by La³⁺, Gd³⁺, Lu³⁺, Sc³⁺ and Y³⁺, of which Gd³⁺ has the greatest enhancement. The experiments indicate that under the optimum condition, the fluorescence intensity of the system is in proportion to the concentration of nucleic acids in the range from 9×10⁻⁸ to 1×10⁻⁵ g ml⁻¹ for yeast RNA, from 1×10⁻⁷ to 1×10⁻⁵ g ml⁻¹ for fish sperm DNA. The detection limits are 3.2 and 4.1 ng ml⁻¹, respectively. This method has been used satisfactorily for the determination of both synthetic and actual samples. In comparison with most fluorescence method for the determination of nucleic acids, this method has higher sensitivity and stability.     

A direct chemiluminescence method for the determination of nucleic acids using Ru(phen)32+–Ce(IV) system

A direct chemiluminescence method for the determination of nucleic acids has been developed based on the enhancement of nucleic acids on the chemiluminescence light emission of the reaction between Ru(phen)₃ ²⁺(phen = 1,10-phenanthroline) and Ce(IV). Under the optimum conditions, the calibration graphs are linear in the range of 5.0 × 10^–8–5.0 × 10^–5 g/mL for calf thymus DNA, 8.0 × 10^–8–5.0 × 10^–5 g/mL for fish sperm DNA and 1.0 × 10^–7–3.0 × 10^–5 g/mL for yeast RNA, respectively. The limits of detection are 1.0 × 10^–8 g/mL for calf thymus DNA, 3.1 × 10^–8 g/mL for fish sperm DNA and 5.2 × 10^–8 g/mL for yeast RNA, respectively. The final procedure allows the successful determination of calf thymus DNA, fish sperm DNA and yeast RNA in six synthetic samples. This method is simple, rapid and specific. Received: 22 January 1999 / Revised: 29 April 1999 / Accepted: 3 May 1999     

Study on the interaction of nucleic acids with silver nanoparticles—Al(III) by resonance light scattering technique and its analytical application

It is found that Al(III) can further enhance the intensity of resonance light scattering (RLS) of the silver nanoparticles (AgNPs) and nucleic acids system. Based on this, a novel method of determination of nucleic acids is proposed in this paper. Under optimum conditions, there are linear relationships between the enhancing extent of RLS and the concentration of nucleic acids in the range of 1.0 × 10⁻⁹-1.0 × 10⁻⁷ g mL⁻¹, 1.0 × 10⁻⁷-2.0 × 10⁻⁶ g mL⁻¹ for fish sperm DNA (fsDNA), 1.0 × 10⁻⁹-7.0 × 10⁻⁸ g mL⁻¹ for calf thymus DNA (ctDNA) and 1.0 × 10⁻⁹-1.0 × 10⁻⁷ g mL⁻¹ for yeast RNA (yRNA). The detection limits (S/N = 3) of fsDNA, ctDNA and yRNA are 4.1 × 10⁻¹⁰ g mL⁻¹, 4.0 × 10⁻¹⁰ g mL⁻¹ and 4.5 × 10⁻¹⁰ g mL⁻¹, respectively. The studies indicate that the RLS enhancement effect should be ascribed to the formation of AgNPs-Al(III)-DNA aggregations through electrostatic attraction and adsorption bridging action of Al(III). And the sensitivity and stability of the AgNPs-fsDNA system could be enhanced by Al(III).     

Near-infrared fluorimetric determination of nucleic acids by shifting the ion-association equilibrium between tetracarboxy aluminum phthalocyanine and tetra-N-hexadecylpyridiniumyl porphyrin

A new method was developed for determination of micro amounts of nucleic acids based on near-infrared (near-IR) fluorescence recovery, employing a two-reagent system which is composed of an anionic tetracarboxy aluminum phthalocyanine (AlC₄Pc) and a cationic tetra-N-hexadecylpyridiniumyl porphyrin (TC₁₆PyP). The fluorescence of the AlC₄Pc, with the maximum emission wavelength at 701 nm, could be quenched by TC₁₆PyP at its proper concentration, but recovered by adding nucleic acids. Under optimal conditions, the recovered fluorescence is proportional to the concentration of nucleic acids. The calibration graphs are linear over the range of 1-200 ng mL⁻¹ for fish sperm DNA (FS DNA) and 2-400 ng mL⁻¹ for calf thymus DNA (CT DNA). The corresponding detection limits are 0.59 ng mL⁻¹ for FS DNA and 0.82 ng mL⁻¹ for CT DNA, respectively. Four synthetic and three real nucleic acid samples were determined with satisfactory results.     

Chemiluminescence flow-through sensor for pipemidic acid using solid sodium bismuthate as an oxidant

A novel chemiluminscence (CL) flow-through sensor for pipemidic acid is described. It was based on the sensitizing effect of pipemidic acid on the CL oxidation of sulfite by sodium bismuthate in H₂SO₄ media. The solid-phase sodium bismuthate was mechanicially immobilized on the sponge rubber inside of the CL flow cell as CL oxidant. The calibration graph is linear in the range 0.1-10 μg/ml with a detection limit of 6.2×10⁻⁸ g/ml (3σ). A complete analysis could be performed in 1 min with a relative standard deviation (R.S.D.) of 2.5% for 2 μg/ml pipemidic acid (n=8). This method has been successfully applied to determine pipemidic acid in pharmaceutical preparation.     

Near-infrared fluorimetric determination of nucleic acids by shifting the ion-association equilibrium between heptamethylene cyanine and Alcian blue 8GX

A new method based on near-infrared (near-IR) fluorescence recovery, employing a two-reagent system which is composed of an anionic heptamethylene cyanine (HMC) and a polycationic phthalocyanine dye, Alcian blue 8GX, is presented for the determination of nucleic acids. With a maximum excitation wavelength at 766 nm and a maximum emission wavelength at 796 nm, the fluorescence recovery is linear with the concentration of nucleic acids added. Factors including the acidity of the medium, the reaction time, the optimal ratio of the two reagents, as well as the influence of foreign substance were all investigated. Meanwhile, the mechanism of fluorescence recovery was also studied. Under the optimal conditions, the linear ranges of the calibration curves were 10-250 ng ml⁻¹ for calf thymus DNA (CT DNA) and 10-200 ng ml⁻¹ for yeast RNA. The detection limits were 6.8 ng ml⁻¹ for CT DNA and 6.3 ng ml⁻¹ for yeast RNA, respectively. The method has been applied to the analysis of practical samples and the recovery results were satisfactory.     

Fluorescent Complexes of Nucleic Acids/8-Hydroxyquinoline/Lanthanum(III) and the Fluorometry of Nucleic Acids

Abstract

The ternary fluorescent complexes of nucleic acids/8-hydroxyquinoline/ lanthanum (III) were studied. Nucleic acids in the study involve natured and thermally denatured calf thymus DNA, fish sperm DNA and yeast RNA. In the range of pH 8.0–8.4 (controlled by NH₃-NH₄Cl buffer) ternary fluorescent complexes are formed which emit at 485.0 nm for calf thymus DNA and at 480.0 nm for yeast RNA (when excited at 267.0 nm) and emits at 483.0 nm for fish sperm DNA when excited at 265.0 nm. Based on the fluorescence reactions sensitive fluorometric methods for nucleic acids were proposed. Using optimal conditions, the calibration curves were linear in the range of 0.4–3.6 μg˙ml^?1 for calf thymus DNA, 0.4–4.0 μg-ml^?1 for fish sperm DNA and 0.4–4.0 μg˙ml^?1 for yeast RNA, respectively. The limits of determination (3σ) were 0.076 μg˙ml^?1 for calf thymus DNA, 0.068 μg˙ml^?1 for fish sperm DNA and 0.329 μg˙ml^?1 for yeast RNA, respectively. Five synthetic samples were determined with satisfaction.

     

Voltammetric study of the interaction of the ofloxacin–copper complex with DNA, and its analytical application

The electrochemical behavior of the ofloxacin–copper complex, Cu(II)L₂, at a mercury electrode, and the interaction of DNA with the complex have been investigated. The experiments indicate that the electrode reaction of Cu(II)L₂ is an irreversible surface electrochemical reaction and that the reactant is of adsorbed character. In the presence of DNA, the formation of the electrochemically non-active complexes Cu(II)L₂-DNA, results in the decrease of the peak current of Cu(II)L₂. Based on the electrochemical behavior of the Cu(II)L₂ with DNA, binding by electrostatic interaction is suggested and a new method for determining nucleic acid is proposed. Under the optimum conditions, the decrease of the peak current is in proportional to the concentration of nucleic acids in the range from 3 × 10⁻⁸ to 3 × 10⁻⁶ g · mL⁻¹ for calf thymus DNA, from 1.6 × 10⁻⁸ to 9.0 × 10⁻⁷ g · mL⁻¹ for fish sperm DNA, and from 3.3 × 10⁻⁸ to 5.5 × 10⁻⁷ g · mL⁻¹ for yeast RNA. The detection limits are 3.3 × 10⁻⁹, 6.7 × 10⁻⁹ and 8.0 × 10⁻⁹ g · mL⁻¹, respectively. The method exhibits good recovery and high sensitivity in synthetic samples and in real samples.     

Study on the interaction between nucleic acid and Eu3+-oxolinic acid and the determination of nucleic acid using the resonance light scattering technique

At pH 9.75, the resonance light scattering (RLS) intensity of OA–Eu³⁺ system is greatly enhanced by nucleic acid. Based on this phenomenon, a new quantitative method for nucleic acid in aqueous solution has been developed. Under the optimum condition, the enhanced RLS is proportional to the concentration of nucleic acid in the range of 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ g/ml for herring sperm DNA, 8.0 × 10⁻¹⁰ to 1.0 × 10⁻⁶ g/ml for calf thymus DNA and 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ g/ml for yeast RNA, and their detection limits are 0.020, 0.011 and 0.010 ng/ml, respectively. Synthetic samples and actual samples were satisfactorily determined. In addition, the interaction mechanism between nucleic acid and OA–Eu³⁺ is also investigated.     

Electrical detection of deoxyribonucleic acid hybridization based on carbon-nanotubes/nano zirconium dioxide/chitosan-modified electrodes

A novel and sensitive electrochemical DNA biosensor based on nanoparticles ZrO₂ and multi-walled carbon nanotubes (MWNTs) for DNA immobilization and enhanced hybridization detection is described. The MWNTs/nano ZrO₂/chitosan-modified glassy carbon electrode (GCE) was fabricated and oligonucleotides were immobilized to the GCE. The hybridization reaction on the electrode was monitored by differential pulse voltammetry (DPV) analysis using electroactive daunomycin as an indicator. Compared with previous DNA sensors with oligonucleotides directly incorporated on carbon electrodes, this carbon nanotube-based assay with its large surface area and good charge-transport characteristics increased DNA attachment quantity and complementary DNA detection sensitivity. The response signal increases linearly with the increase of the logarithm of the target DNA concentration in the range of 1.49 × 10⁻¹⁰ to 9.32 × 10⁻⁸ mol L⁻¹ with the detection limit of 7.5 × 10⁻¹¹ mol L⁻¹ (S/N = 3). The linear regression equation is I = 32.62 + 3.037 log C_DNA (mol L⁻¹) with a correlation coefficient value of 0.9842. This is the first application of carbon nanotubes combined with nano ZrO₂ to the fabrication of an electrochemical DNA biosensor with a favorable performance for the rapid detection of specific hybridization.     

Spectrofluorimetric determination of nucleic acids as 8-hydroxyquinoline/ yttrium ternary complexes

The formation of nucleic acids/8-hydroxyquinoline/yttrium(III) ternary complexes and their fluorescent properties have been studied. The nucleic acids studied include native and thermally denatured calf thymus DNA, fish sperm DNA and yeast RNA. In the range of pH 7.6–8.5, controlled by NH₃-NH₄C1 buffer, ternary complexes are formed that fluoresce at different wavelengths with different nucleic acids. Based on the fluorescence reactions, sensitive spectrofluorimetric methods for nucleic acids are proposed. In optimal conditions, the calibration curves were linear in the range 0.5–4.0 gml^–1 for calf thymus DNA, 0.5–2.5 g ml^–1 for fish sperm DNA and 0.5–4.0 g ml^–1 for yeast RNA. The limits of determination (3 ) were 0.030 g ml^–1 for calf thymus DNA, 0.020 g ml^–1 for fish sperm DNA and 0.090 g ml^–1 for yeast RNA. Corresponding to the interferences of coexisting substances, six synthetic samples were constructed and the results of determination were satisfactory.     

Determination of Nucleic Acids Using Rivanol as the Fluorescent Probe in the Presence of SDS

ABSTRACT

The quantitative method for nucleic acids using rivanol as the fluorescent probe in the presence of SDS was proposed. Under proper conditions, addition of nucleic acids to a mixture of rivanol and SDS resulted in enhanced fluorescence and spectral shifts of rivanol-SDS system. The calibration graph was linear in the range of 0–10.0 μg/ml for CT DNA and 0–9.0 μg/ml for yeast RNA, the limit of detection was 62 ng/ml for CT DNA and 156 ng/ml for yeast RNA. CT DNA could be determined in the presence of 20%(w/w) yeast RNA.     

A New Polarographic Adsorptive Catalytic Wave for the Determination of Trace Tellurium in Smog Dust and Wheat Flour

Linear-sweep polarography of tellurium in sulfuric acid solution containing methylene blue produces a wave at −0.82 V (vs SCE). In a cathodic sweep, the derivative peak current is directly proportional to the concentration of tellurium over the range 4 × 10⁻⁹to 1 × 10⁻⁷g/ml, and the detection limit is 2 × 10⁻⁹g/ml. The polarographic wave is an adsorptive catalytic hydrogen wave. This method has been applied to the determination of trace amounts of tellurium in smog dust and wheat flour, with satisfactory results.     

Quantitative study of stereospecific binding of monoclonal antibody to anti-benzo(a)pyrene diol epoxide-N-dG adducts by capillary electrophoresis immunoassay

The stereospecific binding of monoclonal antibody (mAb) 8E11 to anti-benzo(a)pyrene diol epoxide (BPDE)-dG adducts in single nucleoside, long oligonucleotide, and genomic DNA were quantitatively evaluated using noncompetitive and competitive capillary electrophoresis (CE) immunoassays. Two single-stranded TMR-BPDE-90mers containing a single anti-BPDE-dG adduct with defined stereochemistry and a fluorescent label at 5′-end were used as fluorescent probes for competitive CE immunoassay. To quantitatively evaluate the binding affinity through competitive CE immunoassays, a series of equations were derived according to the binding stoichiometry. The binding of mAb 8E11 to trans-(+)-anti-BPDE-dG displays strongest affinity (K_b: 3.57 × 10⁸ M⁻¹) among all four investigated anti-BPDE-dG mononucleoside adducts, and the cis-(−)-anti-BPDE-dG displays lowest affinity (K_b: 1.14 ×10⁷ M⁻¹). The binding of monoclonal antibody (mAb) 8E11 to BPDE-dG adducts in long DNA (90mer) preferentially forms the complex with a stoichiometry of 1:1, and that mAb 8E11 displays a slightly higher affinity with trans-(+)-anti-BPDE-90mers (K_b: 6.36 ± 0.54 × 10⁸ M⁻¹) than trans-(−)-anti-BPDE-90mers (K_b: 4.52 ± 0.52 × 10⁸ M⁻¹). The mAb 8E11 also displays high affinity with BPDE-dG adducts in genomic DNA (K_b: 3.74 × 10⁸ M⁻¹), indicating its promising applications for sensitive immuno-detection of BPDE-DNA adducts in genomic DNA.     

A novel method for determination of magnesium in urine and water samples with mercury film-plated carbon paste electrode

A square wave adsorptive stripping voltammetric (SWAdSV) method for the indirect determination of trace amounts of magnesium with thiopentone sodium (TPS) as an electroactive ligand, at carbon paste mercury film electrode (CP-MFE) is proposed. It is observed that the increase of the square wave voltammetric cathodic peak current of TPS, under alkaline conditions, is linear with the increase of Mg concentration. Under optimum experimental conditions viz.; pH 10.75, 3×10⁻⁵ M TPS and 0.05 M phosphate buffer (Na₂HPO₄-NaH₂PO₄), a linear relation in the range 6×10⁻⁹ to 9×10⁻⁸ M Mg²⁺ (0.14-2.16 ppb), at 60 s deposition time, is obtained. The detection limit of Mg²⁺ is 0.14 ppb for 60 s deposition time with the relative standard deviation is 0.5% (n=5). The proposed method was successfully applied to the determination of magnesium in urine and tap water samples with satisfactory results. The data obtained are compared with the standard flame atomic absorption spectrophotometric method (FAAS).     

Highly sensitive and selective detection of biothiols using graphene oxide-based “molecular beacon”-like fluorescent probe

A fluorometric method for quantity analysis of biothiols was developed using a graphene oxide (GO)-based “molecular beacon”-like probe, which consisted of FITC labeled thymine (T)-rich single-stranded DNA (ssDNA), GO and Hg²⁺ ions. The labeled ssDNA containing T–T mismatches would self-hybridize to duplex in the presence of Hg²⁺, which can avoid its adsorption on GO and the fluorescence of this GO-based probe was recovered. The fluorescence of the probe quenched after the addition of biothiols such as glutathione (GSH) and cysteine (Cys) owing to thiol groups can selectively competitive ligation of Hg²⁺ ions with T–T mismatches. In the present work, the GO-based probe was used for the determination of GSH and Cys. Under the optimal conditions, a linear correlation was established between fluorescence intensity ratio I₀/I and the concentration of GSH in the range of 2.0 × 10⁻⁹–5.0 × 10⁻⁷ mol L⁻¹ with a detection limit of 1.0 × 10⁻⁹ mol L⁻¹. The linear range for Cys is from 5.0 × 10⁻⁹ to 4.5 × 10⁻⁷ mol L⁻¹ with a detection limit of 2.0 × 10⁻⁹ mol L⁻¹. The proposed method was applied to the determination of GSH in human serum and cell extract samples with satisfactory results.     

Determination of noradrenaline and dopamine in pharmaceutical injection samples by inhibition flow injection electrochemiluminescence of ruthenium complexes

Two maximal potential-resolved flow injection-electrochemiluminescent (FI-ECL) peaks were observed for Ru(bpy)₃²⁺/TPrA system at 0.90 and 1.05 V, and for Ru(phen)₃²⁺/TPrA at 1.01 and 1.25 V (vs. Ag/AgCl) in pH 8.0 phosphate buffer solutions. Sensitive ECL inhibition effects were observed in the presence of noradrenaline and dopamine for both of these systems. Therefore, an FI-ECL inhibition method for determination of noradrenaline and dopamine has been developed. Under optimal conditions, linear responses between logarithm of ECL intensity changes and logarithm of sample concentration were found for noradrenaline in the linear range (LR) of 4×10⁻⁸-1×10⁻⁵ mol l⁻¹ with theoretical detection limit (DL) of 2.5×10⁻⁸ mol l⁻¹ for Ru(bpy)₃²⁺/TPrA system, and in LR of 2×10⁻⁸-2×10⁻⁵ mol l⁻¹ with DL of 7.1×10⁻⁹ mol l⁻¹ for Ru(phen)₃²⁺/TPrA system; and for dopamine in LR of 8×10⁻⁸-2×10⁻⁵ mol l⁻¹ with DL of 5.2×10⁻⁸ mol l⁻¹ for Ru(bpy)₃²⁺/TPrA system, in LR of 4×10⁻⁸-2×10⁻⁵ mol l⁻¹ with DL of 1.5×10⁻⁸ mol l⁻¹ for Ru(phen)₃²⁺/TPrA system. It was applied for determination of commercial pharmaceutical injection samples with satisfied results. The mechanism of the inhibition effects was proposed in the preliminary way.     

Flow injection kinetic spectrophotometric method for the determination of trace amounts of nitrite

In this work, a new, simple and sensitive flow injection catalytic kinetic spectrophotometric determination of nitrite is reported based on catalytic effect of nitrite on the redox reaction between sulfonazo III and potassium bromate in acidic media. The reaction was monitored by measuring the decrease in the absorbance of sulfunazo III at 570 nm. Various chemical (such as the effect of acidity, reagents concentrations) and instrumental parameters (flow rate, reaction coil length, injection volume and temperature) were studied and were optimized. Under the optimum conditions calibration graph was linear in the nitrite concentration ranges of 8.00 × 10⁻³-3.00 × 10⁻¹ μg/ml (with slope of 2.40) and 3.50 × 10⁻¹-1.80 μg/ml (with slope of 0.42). The detection limit was 6.00 × 10⁻³ μg/ml of nitrite, the relative standard deviation (n = 10) was 1.25% and 0.88% for 5.00 × 10⁻² and 2.00 × 10⁻¹ μg/ml of nitrite respectively. About 60 samples in 1 h can be analyzed. The interfering effects of various chemical species were studied. The method was successfully applied in the determination of nitrite in food and environmental samples.     

Fluorometric method for the determination of hydrogen peroxide and glucose with Fe3O4 as catalyst

In this paper, we utilized the instinct peroxidase-like property of Fe₃O₄ magnetic nanoparticles (MNPs) to establish a new fluorometric method for determination of hydrogen peroxide and glucose. In the presence of Fe₃O₄ MNPs as peroxidase mimetic catalyst, H₂O₂ was decomposed into radical that could quench the fluorescence of CdTe QDs more efficiently and rapidly. Then the oxidization of glucose by glucose oxidase was coupled with the fluorescence quenching of CdTe QDs by H₂O₂ producer with Fe₃O₄ MNPs catalyst, which can be used to detect glucose. Under the optimal reaction conditions, a linear correlation was established between fluorescence intensity ratio I₀/I and concentration of H₂O₂ from 1.8 × 10⁻⁷ to 9 × 10⁻⁴ mol/L with a detection limit of 1.8 × 10⁻⁸ mol/L. And a linear correlation was established between fluorescence intensity ratio I₀/I and concentration of glucose from 1.6 × 10⁻⁶ to 1.6 × 10⁻⁴ mol/L with a detection limit of 1.0 × 10⁻⁶ mol/L. The proposed method was applied to the determination of glucose in human serum samples with satisfactory results.     

Electrochemiluminescence of CdTe quantum dots as labels at nanoporous gold leaf electrodes for ultrasensitive DNA analysis

A new electrochemiluminescence (ECL) DNA assay is developed using quantum dots (QDs) as DNA labels. When nanoporous gold leaf (NPGL) electrodes are used, sensitivity of the ECL assay is remarkably increased due to ultra-thin nanopores. In this assay, target DNA (t-DNA) is hybridized with capture DNA (c-DNA) bound on the NPGL electrode, which is fabricated by conjugating amino-modified c-DNA to thioglycolic acid (TGA) modified at the activated NPGL electrode. Following that, amino-modified probe DNA is hybridized with the t-DNA, yielding sandwich hybrids on the NPGL electrode. Then, mercaptopropionic acid-capped CdTe QDs are labeled to the amino group end of the sandwich hybrids. Finally, in the presence of S₂O₈²⁻ as coreactant, ECL emission of the QD-labeled DNA hybrids on the NPGL electrode is measured by scanning the potential from 0 to −2 V to record the curve of ECL intensity versus potential. The maximum ECL intensity (I_m,ECL) on the curve is proportional to t-DNA concentration with a linear range of 5 × 10⁻¹⁵ to 1 × 10⁻¹¹ mol/L. The ECL DNA assay can be used to determine DNA corresponding to mRNA in cell extracts in this study.