Preparation of Luminol-doped Nanoparticle and Its Application in DNA Hybridization Analysis

Introduction The analysis of DNA sequence and DNA mutant detection play fundamental roles in the rapid development of molecular diagnostics and in the anticancer drug screening. Therefor many detection techniques of DNA sequence have been developed in recent years. These techniques mainly depend on the nucleic acid hybridization1 and their sensitivities are related to the specific activity of the label linked to the DNA probe. The degree of hybridization of probe to its complementary DN…     

A Highly Sensitive Electrochemiluminescence Choline Biosensor Based on Poly(aniline‐luminol‐hemin) Nanocomposites

Poly(aniline‐luminol‐hemin) nanocomposites are prepared on an electrode surface through electropolymerization, and a highly sensitive electrochemiluminescence (ECL) biosensor for choline is developed based on the poly(aniline‐luminol‐hemin) nanocomposites and an enzyme catalyzed reaction of choline oxidase (CHOD). The obtained nanocomposites are characterized by scanning electron microscopy (SEM), atomic absorption spectrometry (AAS) and ECL. The results indicate that hemin can be incorporated into the poly(aniline‐luminol) nanocomposites using the facile electropolymerization method, and the poly(aniline‐luminol‐hemin) nanocomposites are rod shaped porous nanostructure. Moreover, the poly(aniline‐luminol‐hemin) nanocomposites exhibit higher ECL intensity than poly(aniline‐luminol) nanocomposites in alkaline media due to the catalytic effect of hemin on the ECL of the polymerized luminol and the electron transfer ability of hemin in the nanocomposites. CHOD is immobilized on the surface of the poly(aniline‐luminol‐hemin) nanocomposites modified electrode with glutaraldehyde, and the ECL biosensor based on poly(aniline‐luminol‐hemin)/CHOD exhibits a wider linear range for the choline detection. The enhanced ECL signals are linear with the logarithm of concentration of choline over the range of 1.0×10^?11～1.0×10^?7 mol L^?1 with a low detection limit of 1.2×10^?12 mol L^?1. Moreover, the proposed biosensor is successfully applied to the detection of choline in milk.     

银（III）配合物-鲁米诺流动注射化学发光新体系测定肾上腺素

儿茶酚胺是一类非常重要的神经递质,在人体的心血管系统、神经系统、内分泌腺、肾脏、平滑肌等组织系统的生理活动中起着广泛的调节作用。肾上腺素为儿茶酚胺的一种,建立灵敏、高效的肾上腺素检测技术具有重要的临床意义。本文将银（Ⅲ）配合物与鲁米诺组成新的流动注射化学发光体系,利用碱性介质中肾上腺素对三价银配合物－鲁米诺化学发光体系有明显的增强效应来测定肾上腺素的含量,并据此建立了高效测定肾上腺素的流动注射化学发光新方法。在优化的条件下,该方法测定肾上腺素的线型范围为1.0×10－9～1.0×10－7 mol L-1,检出限为8.0×10－10 mol L-1,对1.5×10－8 mol L-1肾上腺素11次平行测定,其相对标偏差为2.9％。利用建立的分析方法测定了药物肾上腺素,并对三价银－鲁米诺化学发光新体系测定肾上腺素的反应机理进行了讨论。     

Fabrication of a Sensitive Electrochemical Biosensor for Detection of DNA Hybridization Based on Gold Nanoparticles/CuO Nanospindles Modified Glassy Carbon Electrode

In this work, a sensitive electrochemical DNA biosensor for the detection of sequence‐specific target DNA was reported. Firstly, CuO nanospindles (CuO NS) were immobilized on the surface of a glassy carbon electrode (GCE). Subsequently, gold nanoparticles (Au NPs) were introduced to the surface of CuO NS by the electrochemical deposition mode. Probe DNA with SH (HS‐DNA) at the 5′‐phosphate end was covalently immobilized on the surface of the Au NPs through Au? S bond. Scanning electron microscopy (SEM) was used to elucidate the morphology of the assembled film, and electrochemical impedance spectroscopy technique (EIS) was used to investigate the DNA sensor assembly process. Hybridization detection of DNA was performed with differential pulse voltammetry (DPV) and the methylene blue (MB) was hybridization indicator. Under the optimal conditions, the decline of reduction peak current of MB (ΔI) was linear with the logarithm of the concentration of complementary DNA from 1.0×10^?13 to 1.0×10^?6 mol·L^?1 with a detection limit of 3.5×10^?14 mol·L^?1 (S/N=3). In addition, this DNA biosensor has good selectivity, and even can distinguish single‐mismatched target DNA.     

Detection of DNA immobilized on bare gold electrodes and gold nanoparticle-modified electrodes via electrogenerated chemiluminescence using a ruthenium complex as a tag

Electrogenerated chemiluminescence (ECL) for DNA hybridization detection is demonstrated based on DNA that was self-assembled onto a bare gold electrode and onto a gold nanoparticles modified gold electrode. A ruthenium complex served as an ECL tag. Gold nanoparticles were self-assembled on a gold electrode associated with a 1,6-hexanedithiol monolayer. The surface density of single stranded DNA (ssDNA) on the gold nanoparticle modified gold electrode was 4.8?×?10¹⁴ molecules per square centimeter which was 12-fold higher than that on the bare gold electrode. Hybridization was induced by exposure of the target ssDNA gold electrode to the solution of ECL probe consisting of complementary ssDNA tagged with ruthenium complex. The detection limit of target ssDNA on a gold nanoparticle modified gold electrode (6.7?×?10^?12 mol L^?1) is much lower than that on a bare gold electrode (1.2?×?10^?10 mol L^?1). The method has been applied to the detection of the DNA sequence related to cystic fibrosis. This work demonstrates that employment of gold nanoparticles self-assembled on a gold electrode is a promising strategy for the enhancement of the sensitivity of ECL detection of DNA.     

Electrochemiluminescence Bioassays with a Water‐Soluble Luminol Derivative Can Outperform Fluorescence Assays

The most efficient and commonly used electrochemiluminescence (ECL) emitters are luminol, [Ru(bpy)₃]²⁺, and derivatives thereof. Luminol stands out due to its low excitation potential, but applications are limited by its insolubility under physiological conditions. The water‐soluble m‐carboxy luminol was synthesized in 15 % yield and exhibited high solubility under physiological conditions and afforded a four‐fold ECL signal increase (vs. luminol). Entrapment in DNA‐tagged liposomes enabled a DNA assay with a detection limit of 3.2 pmol L^?1, which is 150 times lower than the corresponding fluorescence approach. This remarkable sensitivity gain and the low excitation potential establish m‐carboxy luminol as a superior ECL probe with direct relevance to chemiluminescence and enzymatic bioanalytical approaches.     

Highly sensitive electrochemiluminescence “turn-on” aptamer sensor for lead(II) ion based on the formation of a G-quadruplex on a graphene and gold nanoparticles modified electrode

We have developed a “turn on” model of an electrochemiluminescence (ECL) based assay for lead ions. It is based on the formation of a G-quadruplex from an aptamer labeled with quantum dots (QDs) and placed on an electrode modified with of graphene and gold nanoparticles (AuNPs). A hairpin capture probe was labeled with a thiol group at the 5′-end and with an amino group at the 3′-end. It was then self-assembled on the electrode modified with graphene and AuNPs. In the absence of Pb(II), the amino tag on one end of the hairpin probe is close to the surface of the electrode and therefore unable to interact with the QDs because of steric hindrance. The ECL signal is quite weak in this case. If, however, Pb(II) is added, the stem-loop of the aptamer unfolds to form a G-quadruplex. The amino group at the 3′-end will become exposed and can covalently link to a carboxy group on the surface of the CdTe QDs. This leads to strong ECL. Its intensity increases (“turns on”) with the concentration of Pb(II). Such a “turn-on” method does not suffer from the drawbacks of “turn-off” methods. ECL intensity is linearly related to the concentration of Pb(II) in the 10 p mol·L^?1 to 1 n mol·L^?1 range, with a 3.8 p mol·L^?1 detection limit. The sensor exhibits very low detection limits, good selectivity, satisfying stability, and good repeatability.

A lipase-based electrochemical biosensor for target DNA

A lipase-based electrochemical biosensor has been fabricated for the quantitative determination of target DNA. It is based on a stem-loop nucleic acid probe labeled with ferrocene containing a butanoate ester that is hydrolyzed by lipase. The other end of the probe DNA is linked, via carboxy groups, to magnetic nanoparticles. The binding of target DNA transforms the hairpin structure of the probe DNA and causes the exposure of ester bonds. This results in the release of electro-active ferrocene after hydrolysis of the ester bonds, and in an observable electrochemical response. The quantity of target DNA in the concentration range between 1?×?10^?12 mol·L^?1 and 1?×?10^?8 mol·L^?1 can be determined by measuring the electrochemical current. The method can detect target DNA with rapid response (30 min) and low interference.

Sensitive Nonenzymatic Electrochemiluminescence Determination of Hydrogen Peroxide in Dental Products using a Polypyrrole/Polyluminol/Titanium Dioxide Nanocomposite

A layer-by-layer assembled of a polypyrrole and polyluminol was synthesized through the electrodeposition of pyrrole and luminol in acidic medium on a graphite electrode. The electrode was then modified by casting titanium dioxide (TiO₂) nanoparticles on its surface for enhancing electrochemiluminescence of luminol. The properties of this electrochemiluminescence sensor were studied by cyclic voltammetry, electrochemical impedance spectroscopy, field emission scanning electron microscopy, and energy dispersive X-ray spectroscopy. The results demonstrated that the modification of this electrochemiluminescence sensor shows sensitive response for the determination of hydrogen peroxide. Figures of merit include broad linearity from 1?pmol L^?1 to 4?µmol L^?1 (R²?=?0.996) with a limit of detection as low as 0.40?pmol L^?1 at a signal-to-noise ratio of three and good reproducibility with relative standard deviation of 4% for the determination of a 400?nmol L^?1 hydrogen peroxide solution (n?=?4), along with favorable long-term stability. The presence of glucose, citric acid, uric acid, dopamine, and ascorbic acid at concentrations as high as 100?nmol L^?1 of H₂O₂ did not produce any electrochemiluminescence signals, which demonstrates the selective nature of this modified electrode. The sensor was also used for the determination of H₂O₂ in mouthwash formulations and dental whitelight gels.     

Highly Sensitive Electrochemiluminescence Detection of Mercury(II) Ions Based on DNA‐Linked Luminol‐Au NPs Superstructure

A novel highly sensitive electrochemiluminescence (ECL) detection protocol for mercury(II) ions was developed. Based on the strong and stable thymine? thymine mismatches complexes coordination chemistry, mercury(II) ions can specifically bind to a designed DNA strand, leading to the release of the complimentary DNA strand. The released DNA strand was then captured by magnetic beads modified with specific DNA, and then through the formation of DNA‐linked luminol‐Au nanoparticles (NPs) superstructure, a specific ECL system for mercury(II) ions was developed. Using 3‐aminopropyl‐triethoxysilane as an effective enhancer, the ECL system can detect Hg²⁺ ion within a linear range from 2.0×10^?10 mol L^?1 to 2.0×10^?8 M, with a detection limit as low as 1.05×10^?10 M (3σ). Moreover, this ECL system is highly specific for Hg²⁺, without interference from other commonly coexisted metal ions, and it can be used for the analysis of real samples.     

Chitosan‐Based Glucose Oxidase Electrodes Enhanced by Silver Nanoparticles

Silver nanoparticles enhanced glucose oxidase electrodes were prepared on the basis of chitosan matrix. The enzyme electrodes exhibited high sensitivity and excellent response performance to glucose with a linear range from 1×10^?6 to 8×10^?3 mol · L^?1. And the time reaching the steady‐state amperometric response was less than 5 seconds. The inhibition percentage of this enzyme electrode against copper ions concentration was linear ranging from 1.2×10^?6 to 5×10^?5 mol · L^?1. These properties of enzyme electrodes are probably due to the excellent electron transfer of silver nanoparticles and the orientation of glucose oxidase molecule.     

Alizarin-modified sulfonate carbon nanoparticles in vanadium sensing

The adsorption processes of alizarin onto hydrophilic carbon nanoparticles (Emperor 2000?) are investigated. The significant increase in voltammetric responses for pre-adsorbed alizarin compared with those for solution confirms high affinity of alizarin to carbon nanoparticles (possibly due to π–π stacking interaction between aromatic rings of alizarin and surface-sulfonated carbon nanoparticles). To obtain the optimum of adsorption conditions, the effects of pH, agitation rate, and adsorption time are investigated. Under square wave voltammetry conditions, the peak current for the reduction of alizarin shows a linear relationship with concentration in the range from 2.0 to 10.0 nM. The limit of detection is estimated 5.8?×?10^?9 mol L^?1. Next, alizarin is applied as a receptor for sensing of trace vanadium in acetate buffer pH 5. Linear calibration curves are obtained for vanadium in the range of 1.0?×?10^?6 to 1.0?×?10^?4 mol L^?1 and the limit of detection is estimated 9.6?×?10^?8 mol L^?1. Determination of vanadium in real samples such as sea and tap water is demonstrated.     

Determination of Hydrogen Peroxide Using a Novel Sensor Based on Fe3O4 Magnetic Nanoparticles

Fe₃O₄ magnetic nanoparticles were synthesized by chemical co-precipitation with sodium citrate as a surfactant and were used with chitosan to construct a novel hydrogen peroxide sensor. The electrochemical behavior of hydrogen peroxide at the sensor was investigated by cyclic voltammetry. The composite film electrocatalyzed the reduction of hydrogen peroxide, and the peak current increased linearly with concentration from 1.00 × 10^?5 to 1.00 × 10^?3 mol · L^?1 (R = 0.9974) with a detection limit of 1.53 × 10^?6 mol · L^?1. This novel nonenzyme sensor provided good sensitivity, stability, and precision with potential applications.     

L-Argininamide biosensor based on S1 nuclease hydrolysis signal amplification

A sensitive method is presented for the detection of L-argininamide. It is based on the amplification of the hydrolysis of S1 nuclease of single-stranded regions of an aptamer-target complex. The S1 nuclease, which is sequence-independent, is used to “recycle” target molecules, thus leading to strongly enhanced chemiluminescence (CL). L-Argininamide was chosen as model analyte. The DNA aptamer and its complementary DNA were labeled with the CL reagent N-(4-aminobutyl)-N-ethylisoluminol (ABEI). The DNA complementary to the aptamer was labeled with ABEI and immobilized on magnetic beads (MBs) coated with gold. The aptamer was also labeled with ABEI and self-assembled on the MBs. A duplex was formed due to hybridization between the DNA aptamer and the DNA complementary to the aptamer. In the presence of the target L-argininamide, a stem-loop aptamer structure is formed which subsequently denatures the duplex. This switch from a duplex structure to a stem-loop structure causes the formation of single-stranded regions both in the target-aptamer and in the single-stranded DNA on the MBs. The nuclease hydrolyzes the single-stranded regions and single-stranded DNA. Ultimately, L-argininamide is released which then interacts with another aptamer on the MB, thereby leading to one more L-argininamide. This autocatalytic cycle can generate substantial quantities of ABEI which then can be sensitively determined by the diperiodatonickelate-isoniazide reaction system. L-argininamide can be detected in the concentration range from 3.0?×?10^?4 to 3.0?×?10^?7 M, and the limit of detection is 1.0?×?10^?7 M.

Highly Sensitive Electrogenerated Chemiluminescence Isoniazid with NiO Nanoparticles‐modified Graphite Electrode

Abstract

NiO nanoparticles (NiO NPs) were prepared with chemical precipitation method and modified on the surface of vaseline‐impregnated graphite electrode with chitosan. It was found that, based on the catalysis of the NiO NPs for the chemiluminescent reaction of the ECL process, the enhancing effect of isoniazid on the weak electrogenerated chemiluminescence (ECL) signal of luminol at a NiO NPs‐chitosan modified electrode was stronger than that at a bare graphite electrode. Under the optimum experimental conditions, the relative ECL intensity was linear with isoniazid concentration over the range 3.0×10^?10～1.0×10^?6 g/ml at the NiO NPs‐chitosan modified electrode with a detection limit of 1.0×10^?10 g/ml.     

Fabrication of a Sensitive Impedance Biosensor of DNA Hybridization Based on Gold Nanoparticles Modified Gold Electrode

In this work, we report on the preparation of a simple, sensitive DNA impedance sensor. Firstly gold nanoparticles were electrodeposited on the surface of a gold electrode, and then probe DNA was immobilized on the surface of gold nanoparticles through a 5′‐thiol‐linker. Electrochemical impedance spectroscopy (EIS) was used to investigate probe DNA immobilization and hybridization. Compared to the bare gold electrode, the gold nanoparticles modified electrode could improve the density of probe DNA attachment and the sensitivity of DNA sensor greatly. The difference of electron transfer resistance (ΔR_et) was linear with the logarithm of complementary oligonucleotides sequence concentrations in the range of 2.0×10^?12 to 9.0×10^?8 M, and the detection limit was 6.7×10^?13 M. In addition, the DNA sensor showed a fairly good reproducibility and stability during repeated regeneration and hybridization cycles.     

Electrochemical DNA biosensor for the detection of Trichoderma harzianum based on a gold electrode modified with a composite membrane made from an ionic liquid, ZnO nanoparticles and chitosan, and by using acridine orange as a redox indicator

An electrochemical DNA biosensor was developed that is based on a gold electrode modified with a nanocomposite membrane made from an ionic liquid, ZnO nanoparticles and chitosan. A single-stranded DNA probe was immobilized on this electrode. Acridine orange was used as the hybridization probe for monitoring the hybridization of the target DNA. The biosensor was capable of detecting target DNA in the concentration range from 1.0?×?10?C14 to 1.8?×?10?C4?mol?L-1, with a detection limit of 1.0?×?10?C15?mol?L-1. The approach towards constructing a DNA biosensor allows studies on the hybridization even with crude DNA fragments and also to analyze sample obtained from real samples. The results show that the DNA biosensor has the potential for sensitive detection of a specific sequence of the Trichoderma harzianum gene and provides a quick, sensitive and convenient method for the study of microorganisms.

健那绿B的荧光猝灭研究及作为核酸测定探针的应用

用紫外可见光谱、稳态荧光发射及荧光寿命测定研究了核酸猝灭十二烷基磺酸钠胶束中的健那绿荧光。水溶液中弱的健那绿荧光在十二烷基磺酸钠胶束中被大大加强，其最大发射从425纳米移至410纳米，核酸的加入将猝灭健那绿的荧光，当健那绿浓度为2.5×10^–5 mol•L^-1时，荧光猝灭(F₀/F)分别与小牛胸腺DNA及鱼精DNA在2.4×10^–8 到 1.08×10^–7及 1.9×10^–8 到 3.8×10^–8 mol•L^-1范围内成正比, 检测限分别为1.3×10^–8 mol•L^-1 (小牛胸腺DNA)及6.3×10^–9 mol•L^-1 (鱼精DNA)。当DNA浓度较高时, 将系统偏离Stern-Volmer方程。这是因为动态猝灭和静态猝灭同时存在。方法已应用于鸡血提取液中DNA的测定, 测定结果与紫外法一致。     

Magnetic nanoparticles grafted with β-cyclodextrin for solid-phase extraction of 5-hydroxy-3-indole acetic acid

We describe the synthesis of ß-cyclodextrin modified magnetic nanoparticles (CD-mNPs) as a material for solid-phase extraction of the cancer biomarker 5-hydroxy-indole-3-acetic acid (5-HIAA) from urine. The CD-mNPs were characterized by TEM, FTIR, and XRD, and the kinetics and adsorption isotherms were studied. The strong interaction between the CD-mNPs and 5-HIAA is the main driving force for recognition and extraction, while the magnetic core of the NPs allows their separation from the sample matrix. Recovery of 5-HIAA from the adsorbent using an adequate solvent regenerated the adsorbent for further use. 5-HIAA was then quantified by fluorometry of its complex with ß-CD. The method works in the 1?×?10^?7 to 1?×?10^?5 mol L^?1 (R² 0.9982–0.9996) concentration range, and the limits of detection (3σ) and quantification (10 σ) of the method are 1.2?×?10^?8 mol L^?1 and 4.01?×?10^?8 mol L^?1 5-HIAA, respectively. The recovery of 5-HIAA from urine samples spiked with 5-HIAA in three concentrations (1.4?×?10^?6, 4.50?×?10^?6 and 1.0?×?10^?5 mol L^?1) are within 63?±?3 %.

Molecular imprinting polymer electrosensor based on gold nanoparticles for theophylline recognition and determination

An electrochemical sensor for theophylline (ThPh) was prepared by electropolymerizing o-phenylenediamine on a glassy carbon electrode in the presence of ThPh via cyclic voltammetry, followed by deposition of gold nanoparticles using a potentiostatic method. The effects of pH, ratio between template molecule and monomer, number of cycles for electropolymerization, and of the solution for extraction were optimized. The current of the electro-active model system hexacyanoferrate(III) and hexacyanoferrate(IV) decreased linearly with successive addition of ThPh in the concentration range between 4.0?×?10^?7?~?1.5?×?10^?5 mol·L^?1 and 2.4?×?10^?4?~?3.4?×?10^?3 mol·L^?1, with a detection limit of 1.0?×?10^?7 mol·L^?1. The sensor has an excellent recognition capability for ThPh compared to structurally related molecules, can be regenerated and is stable.