Double‐Layer Nanogold and Double‐Strand DNA‐Modified Electrode for Electrochemical Immunoassay of Cancer Antigen 15‐3

Various sensor‐based immunoassay methods have been extensively developed for the detection of cancer antigen 15‐3 (CA 15‐3), but most often exhibit low detection signals and low detection sensitivity, and are unsuitable for routine use. The aim of this work is to develop a simple and sensitive electrochemical immunoassay for CA 15‐3 in human serum by using nanogold and DNA‐modified immunosensors. Prussian blue (PB), as a good mediator, was initially electrodeposited on a gold electrode surface, then double‐layer nanogold particles and double‐strand DNA (dsDNA) with the sandwich‐type architecture were constructed on the PB‐modified surface in turn, and then anti‐CA 15‐3 antibodies were adsorbed onto the surface of nanogold particles. The double‐layer nanogold particles provided a good microenvironment for the immobilization of biomolecules. The presence of dsDNA enhanced the surface coverage of protein, and improved the sensitivity of the immunosensor. The performance and factors influencing the performance of the immunosensor were evaluated. Under optimal conditions, the proposed immunosensor exhibited a wide linear range from 1.0 to 240 ng/mL with a relatively low detection limit of 0.6 ng/mL (S/N=3) towards CA 15‐3. The stability, reproducibility and precision of the as‐prepared immunosensor were acceptable. 57 serum specimens were assayed by the developed immunosensor and standard enzyme‐linked immunosorbent assay (ELISA), respectively, and the results obtained were almost consistent. More importantly, the proposed methodology could be further developed for the immobilization of other proteins and biocompounds.     

Graphene and Nanogold‐Functionalized Immunosensing Interface with Enhanced Sensitivity for One‐Step Electrochemical Immunoassay of Alpha‐Fetoprotein in Human Serum

A new electrochemical immunosensing protocol for sensitive detection of alpha‐fetoprotein (AFP, as a model) in human serum was developed by means of immobilization of horseradish peroxidase‐anti‐AFP conjugates (HRP‐anti‐AFP) onto graphene and nanogold‐functionalized biomimetic interfaces. The low‐toxic and high‐conductive graphene complex provided a large capacity for nanoparticulate immobilization and a facile pathway for electron transfer. With a one‐step immunoassay format, the antigen‐antibody complex was formed between the immobilized HRP‐anti‐AFP on the electrode and AFP in the sample. The formed immunocomplex was coated on the electrode surface, inhibited partly the active center of HRP, and decreased the catalytic reduction of HRP toward the enzyme substrate of H₂O₂. Under optimal conditions, the decrease of reduction currents was proportional to AFP concentration, and the dynamic range was 1.0–10 ng/mL with a relative‐low detection limit (LOD) of 0.7 ng/mL AFP. Intra‐ and inter‐assay coefficients of variation (CVs) were less than 10 %. The assay was evaluated for clinical human serum samples, including 8 (possible) patients with hepatocarcinoma and 3 normal human sera. Correct identification of negative/positive samples and perfect accordance with results from Elecsys 2010 Electrochemiluminescent Automatic Analyzer as a reference was obtained. Importantly, the graphene and nanogold‐based sensor provided a promising platform for the detection of other biocompounds, and could be further applied for development of other potential electrochemical bio/chemosensors.     

Electrochemical Immunoassay for Free Prostate Specific Antigen (f‐PSA) Using Magnetic Beads

Prostate specific antigen (PSA) is a prominent marker for the prostate carcinoma. It is found in human blood in free (f‐PSA) and complex forms. These two forms together are called total PSA (t‐PSA). Estimation of both forms is essential to predict malignancy. In this study we report a unique and effective technique of electrochemical detection of f‐PSA using magnetic beads on a three‐electrode screen‐printed sensor. A magnetic bead enzyme linked immunosorbent assay (ELISA) was performed in a cuvette. Following the immunoassay, magnetic beads were recovered by a magnetic concentrator and transferred on the working electrode of the 3‐electrode assembly. The amperometric response, a measure of the amount of residual enzyme activity on the beads and hence the concentration of analyte in solution, was determined by addition of enzyme substrate. The device has a detection limit of <0.1 ng mL^?1 f‐PSA and a linear range of 0 to 1 ng mL^?1 f‐PSA.     

Nanogold-penetrated poly(amidoamine) dendrimer for enzyme-free electrochemical immunoassay of cardiac biomarker using cathodic stripping voltammetric method

Methods based on immunoassays have been developed for cardiac biomarkers, but most involve the low sensitivity and are unsuitable for early disease diagnosis. Herein we design an electrochemical immunoassay for sensitive detection of myoglobin (a cardiac biomarker for acute myocardial infarction) by using nanogold-penetrated poly(amidoamine) dendrimer (AuNP-PAMAM) for signal amplification without the need of natural enzymes. The assay was carried out on the monoclonal mouse anti-myoglobin (capture) antibody-anchored glassy carbon electrode using polyclonal rabbit anti-myoglobin (detection) antibody-labeled AuNP-PAMAM as the signal tag. In the presence of target myoglobin, the sandwiched immunocomplex could be formed between capture antibody and detection antibody. Accompanying AuNP-PAMAM, the carried gold nanoparticles could be directly determined via stripping voltammetric method under acidic conditions. Under optimal conditions, the detectable electrochemical signal increased with the increasing target myoglobin in the sample within a dynamic working range from 0.01 to 500 ng mL⁻¹ with a detection limit of 3.8 pg mL⁻¹. The electrochemical immunoassay also exhibited high specificity and good precision toward target myoglobin. Importantly, our strategy could be applied for quantitative monitoring of myoglobin in human serum specimens, giving well matched results with those obtained from commercialized enzyme-linked immunosorbent assay (ELISA) method.     

A Novel Inorganic-Organic Composite Constructed from Cobalt（Ⅱ）-Monosubstituted Polyoxometalates and Poly（amidoamine）-NH2 DendrimersA Novel Inorganic-Organic Composite Constructed from Cobalt（Ⅱ）-Monosubstituted Polyoxometalates and Poly（amidoamine）-NH2 Dendrimers

An inorganic-organic composite was prepared by poly（amidoamine） （PAMAM） dendrimer reacting with cobalt（Ⅱ）-monosubstituted polyoxometalates Na5Co^Ⅱ（H2O）PW11O39 （PW11CO） in an aqueous solution. The hybrid composite PW11Co/PAMAM was characterized by FT-IR, UV-Vis diffuse reflectance spectra （DR-UV-Vis）, XPS, XRD and TG/DTA, indicating that the PWI iCo was chemically anchored to PAMAM. The morphologies of the title composite were characterized by scanning electron microscopy （SEM） and transmission electron microscopy （TEM）. The catalytic activity was evaluated by oxidation of isobutyraldehyde （IBA） to isobutyric acid （IBAc） in MeCN under mild conditions （20 ℃, ambient pressure）, showing that the title compound is a more effective and recoverable catalyst than corresponding PW11Co.     

Poly(amidoamine) dendrimer‐coated magnetic nanoparticles for the fast purification and selective enrichment of glycopeptides and glycans

Glycosylated proteins modulate various important functions of organisms. To reveal the functions of glycoproteins, in‐depth characterization studies are necessary. Although mass spectrometry is a very efficient tool for glycoproteomic and glycomic studies, efficient sample preparation methods are required prior to analyses. In the study, poly(amidoamine) dendrimer‐coated magnetic nanoparticles were presented for the specific enrichment and fast purification of glycopeptides and glycans. The enrichment and purification performance of the developed method was evaluated both at the glycopeptide, and the glycan level using several standard glycoprotein digests and released glycan samples. The poly(amidoamine) dendrimer‐coated magnetic nanoparticles not only showed selective affinity (Immunoglobulin G/Bovine Serum Albumin, 1/10 by weight) to glycopeptides and released glycans but also good sensitivity (0.4 ng/µL for Immunoglobulin G) for glycoproteomic and glycomic applications. Thirty‐five glycopeptides of Immunoglobulin G were detected after enrichment with poly(amidoamine) dendrimer‐coated magnetic nanoparticles. In addition, 55 ¹⁸O tagged deamidated glycopeptides belonging to human plasma glycoproteome were confirmed. Finally, fifty 2‐aminobenzoic acid, and 30 procainamide‐labelled human plasma N‐glycans released from human plasma glycoproteins were determined after purifications. The results indicate that the proposed enrichment and purification method using poly(amidoamine) dendrimer‐coated magnetic nanoparticles could be simply adjusted to sample preparation methods.     

基于酶标抗体和媒介体共吸附于金胶壳聚糖膜的PSA免疫传感器的制备

本文研制了一种用金胶壳聚糖仿生膜来同时固定四甲基联苯胺（TMB）和酶标抗体的新型电化学免疫传感器,用于检测血清肿瘤标志物前列腺特异性抗原（PSA）的含量。固定的TMB作为电子传递媒介体,在扫速小于45 mV/s时,电极表现为一个表面控制过程,而在扫速大于45 mV/s时则表现为一个扩散控制过程。将固定有酶标抗体和TMB的免疫传感器与待测PSA抗原一起培育,在该传感器上形成的免疫复合物通过TMB-H₂O₂-HRP电化学体系进行了测定。在优化实验条件下,PSA的线性检测范围为5-30 ng·mL^-1,检测限为1.0 ng·mL^-1。该PSA免疫传感器制备方法简单,成本低廉,具有较好的稳定性和重现性。     

Electrochemical Study and Application on Shikonin at Poly(diallyldimethylammonium chloride) Functionalized Graphene Sheets Modified Glass Carbon Electrode

The electrochemical behaviors of shikonin at a poly(diallyldimethylammonium chloride) functionalized graphene sheets modified glass carbon electrode(PDDA-GS/GCE) have been investigated. Shikonin could exhibit a pair of well-defined redox peaks at the PDDA-GS/GCE located at 0.681 V(E_pa) and 0.662 V(E_pc)[vs. saturated calomel electrode(SCE)] in 0.1 mol/L phosphate buffer solution(pH=2.0) with a peak-to-peak separation of about 20 mV, revealing a fast electron-transfer process. Moreover, the current response was remarkably increased at PDDA-GS/GCE compared with that at the bare GCE. The electrochemical behaviors of shikonin at the modified electrode were investigated. And the results indicate that the reaction involves the transfer of two electrons, accompanied by two protons and the electrochemical process is a diffusional-controlled electrode process. The electrochemical parameters of shikonin at the modified electrode, the electron-transfer coefficient(α), the electron-transfer number(n) and the electrode reaction rate constant(k_s) were calculated to be as 0.53, 2.18 and 3.6 s^-1, respectively. Under the optimal conditions, the peak current of differential pulse voltammetry(DPV) increased linearly with the shikonin concentration in a range from 9.472×10^-8 mol/L to 3.789×10^-6 mol/L with a detection limit of 3.157×10^-8 mol/L. The linear regression equation was I_p=0.7366c+0.7855(R=0.9978; I_p: 10^-7 A, c: 10^-8 mol/L). In addition, the modified glass carbon electrode also exhibited good stability, selectivity and acceptable reproducibility that could be used for the sensitive, simple and rapid determination of shikonin in real samples. Therefore, the present work offers a new way to broaden the analytical application of graphene in pharmaceutical analysis.     

Electrochemical Sensor Based on Thioether Oligomer Poly(N‐vinylpyrrolidone)‐modified Gold Electrode for Bisphenol A Detection

Presently, bisphenol A (BPA) has been added to the list of substances of very high concern as endocrine disruptors. According to the literature, exposure to bisphenol A even at low doses may result in adverse health effects. In this study, electrochemical sensor of Bisphenol A based on thioether DDT‐Poly(N‐vinylpyrrolidone) oligomer has been developed. The thioether oligomer, which is capable of recognizing BPA, was prepared and used for gold electrode modification. The characterization of the modified gold electrode and the synthesized thioether oligomer were carried out by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), Fourier‐transform infrared spectroscopy (FTIR), proton nuclear magnetic resonance spectroscopy (¹H NMR) and Size exclusion chromatography (SEC). Obtained results indicate that the modified electrode shows good electrochemical activity, good sensitivity and reproducibility for BPA detection. It exhibited a good linear relationship ranging from 1 to 20 pg/mL, and the detection limit was found to be 1.9 pg/mL at S/N=3. Several interfering species such as hydroquinone, phenol and resorcinol were used and their behaviors on the modified gold electrode were investigated.     

聚萘胺膜与金属离子的络合及催化特性的研究

系统研究了聚萘胺膜与Ａｇ、Ｈｇ（２＋）等十余种金属离子的络合特性。以循环伏安法、交流阻抗分析等方法研究了Ａｇ－ＰＮＡ电极对抗坏血酸（Ｈ２Ａ）氧化的催化作用，提出了电极反应的可能机理。Ａｇ－ＰＮＡ电极可用于测定试样中的Ｈ２Ａ含量。     

Dendritic Carrier Based on PEG: Design and Degradation of Acid‐sensitive Dendrimer‐like Poly(ethylene oxide)s

Degradable dendrimer‐like PEOs were designed using an original ABC‐type branching agent featuring a cleavable ketal group, following an iterative divergent approach based on the anionic ring opening polymerization (AROP) of ethylene oxide and arborization of PEO chain ends. A seventh generation dendrimer‐like PEO carrying 192 peripheral hydroxyls and exhibiting a molar mass of 446 kg · mol⁻¹ was obtained in this way. The chemical degradation of these dendritic scaffolds was next successfully accomplished under acidic conditions, forming linear PEO chains of low molar mass (≈2 kg · mol⁻¹), as monitored by ¹H NMR, SEC, and MALDI‐TOF mass spectrometry as well as by AFM.

     

聚吖啶橙修饰电极的电化学行为及其对肾上腺素的电催化性能

研究了聚吖啶橙 (POAO)修饰电极及其电化学性能 ,并用于肾上腺素 (EP)的电化学测定。EP在POAO修饰电极上产生一灵敏的氧化峰 ,与裸玻碳电极 (GCE)相比 ,其峰电位负移了 2 30mV ,明显降低了EP的氧化过电位。在pH 6 .0的磷酸氢二钠 柠檬酸缓冲溶液中 ,氧化峰电流与EP的浓度在 4 .5× 10 - 7～ 9.2× 10 - 5mol L范围内呈良好的线性关系 ,检出限为 1.0× 10 - 9mol L。可用于实际样品中EP的测定     

Poly(diallydimethylammonium chloride) Functionalized Graphene/Double‐walled Carbon Nanotube Composite for Amperometric Determination of Nitrite

Poly(diallydimethylammonium chloride) functionalized graphene/double‐walled carbon nanotube composite (PDDA‐GN/DWCNT) was synthesized by a facile microwave‐assisted reduction reaction. PDDA‐GN/DWCNT exhibits enhanced electrocatalytic activity towards nitrite oxidation. The direct electrooxidation behaviors of nitrite on PDDA‐GN/DWCNT were investigated by cyclic voltammetry, amperometric i‐t curve, and rotating disk electrode experiments. Under the optimum conditions, PDDA‐GN/DWCNT was applied to the quantitative analysis of nitrite with a wide linear range covering 0.07–3000 µM and a low detection limit of 0.03 µM (S/N=3). Moreover, PDDA‐GN/DWCNT exhibits satisfied results in the determination of nitrite that released from breast cancer cells.     

侧链含有全氟磺酸的聚芳醚砜质子交换膜材料的制备及性能

利用含溴苯侧基的聚芳醚砜(PES-Br)和1,1,2,2-四氟-2-(1,1,2,2-四氟-2-碘乙氧基)乙烷磺酸钾(PSA-K)进行Ullman偶联反应, 制备了新型含全氟磺酸侧链的聚芳醚砜(PES-PSA), 全氟磺酸的引入量为50%. 在离子交换容量(IEC)为0.907 meq./g, 80 ℃, 相对湿度为100%时, PES-PSA质子传导率达到0.039 S/cm, 表现出较高的质子传导能力. 其较低的IEC值使膜材料表现出良好的热稳定性和尺寸稳定性.     

Dual Amplified Electrochemical Immunosensor for Hepatitis B Virus Surface Antigen Detection Using Hemin/G‐Quadruplex Immobilized onto Fe3O4‐AuNPs or (Hemin‐Amino‐rGO‐Au) Nanohybrid

A sensitive electrochemical immunosensor for Hepatitis B virus surface antigen (HBsAg) detection was fabricated based on hemin/G‐quadruplex interlaced onto Fe₃O₄‐AuNPs or hemin ‐amino‐reduced graphene oxide nanocomposite (H‐amino‐rGO‐Au). G‐quadruplex DNAzyme, which is composed of hemin and guanine‐rich nucleic acid, is an effective signal amplified tool for its outstanding peroxidase activity and Fe₃O₄‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites with quasi‐enzyme activity provide appropriate support for the immobilization of hemin/G‐quadruplex. The target protein was sandwiched between the primary antibody immobilized on the GO and secondary antibody immobilized on the Fe₃O₄‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites and glutaraldehyde was used as linking agent for the immobilization of primary antibody on the surface of GO. Both Fe₃O₄‐AuNPs and H‐amino‐rGO‐Au nanocomposite and also hemin/G‐quadruplex can cooperate the electrocatalytic reduction of H₂O₂ in the presence of methylene blue as mediator. The proposed immunosensor has a wide linear dynamic range of 0.1 pg/ml to 300 pg/ml with a detection limit of 60 fg/ml when Fe₃O₄‐AuNPs was used for immobilization of hemin/G‐quadruplex, while the dynamic range and DL were 0. 1–1000 pg/mL and 10 fg/mL, respectively in the presence of H‐amino‐rGO‐ Au nanocomposite as platform for immobilizing of hemin/G‐quadruplex. The proposed immunosensor was also used for analysis of HBsAg in spiked human serum samples with satisfactory results.     

The Influence of the Cathodic Pretreatment on the Electrochemical Detection of Dopamine by Poly(1‐aminoanthracene) Modified Electrode

In this study we demonstrated the influence of the cathodic pretreatment of poly(1‐aminoanthracene) (PAA) electropolymerized on a platinum electrode for determination of dopamine (DA). The DA electrochemical response was obtained after a cathodic pretreatment of the PAA electrode which consisted of applying a potential of ?0.7 V (vs. Ag/AgCl) for 3 s before each measurement. The pretreatment of the electrode changed the PAA electrocatalytic properties so that the electrode began to present electrochemical response to DA without interference of ascorbic acid (AA). The anodic peak currents determined by differential pulse voltammetry using pretreated PAA showed a linear dependence on the DA concentration from 0.56 to 100 µM with a detection limit of 0.13 µM and a correlation coefficient of 0.9986. The electrode exhibits a relative standard deviation of 1.2 % for ten successive measurements of a 0.5 mM DA solution. The analysis by scanning electron microscopy and atomic force microscopy show a homogeneous and nanostructured film with globular structures with diameter of about 20 nm. The analytical results obtained for DA determination at a pretreated PAA electrode in pharmaceutical formulation sample were in good agreement with those obtained by a comparative procedure at a 95 % confidence level. PAA electrode after the pretreatment showed electrochemical responses to DA with excellent selectivity, sensitivity, and high stability without interference of AA.     

Hetero‐difunctional dimers as building blocks for the synthesis of poly(amidoamine)s with hetero‐difunctional chain terminals and their derivatives

This article reports on a simple and straightforward preparation method of poly(amidoamine)s (PAAs) with hetero‐difunctional chain ends as well as of several up to now hardly obtainable PAA derivatives of biotechnological interest, such as for instance PAAs of controlled molecular weight and narrow polydispersity mono‐functionalized at one end with an acrylamide group, PAAs with star‐like molecular architecture, graft‐PAA‐protein conjugates, “tadpole‐like” PAA conjugates with hydrophobic moieties able to self assemble into nanoparticles in aqueous media. The key step was to design suitable building blocks consisting of hetero‐difunctional dimers (HDDs). In particular, the HDDs considered were the mono‐addition products of bis‐sec‐amines and bisacrylamides expected to give PAAs of proven biomedical potential and were obtained as hydrochlorides or trifluoroacetates. In this form, they could be indefinitely kept dormant at 0–5 °C in the dry state, whereas at room temperature and in aqueous media, they polymerized at pH > 7.5. The preparation of the above‐cited PAA derivatives did not necessarily involve the preliminary synthesis of hetero‐difunctional PAAs but was directly achieved by one‐pot polymerization of HDDs in the presence of the substrates of interest. © 2012 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2012     

AC Impedance Studies on the Relationship between the Fractal Characteristics and Electrochemical Properties of Poly(Para‐Phenylene) Film

The effects of experimental conditions on the fractal structure of electrosynthetic polyparaphenylene films were studied by electrochemical impedance spectroscopy. The results indicate that, at the potential range (0‐0.6 V), the film surface fractal dimension rises while the corresponding charge transfer resistances R_ct decrease as charge increases. This is consistent with the oxidized behavior of the conducting polymer. As for the degradation of electrolytes, scanning electron microscopy observations of poly(para‐pheneylene) (PPP) film provide evidence of the close relationship between the degradation of electrolytes and the film morphology, also in good agreement with the electrochemical impedance results interpretation. The X‐ray photoelectron spectroscopic analysis results present that the relative oxygen content ratio (O/C) of the films increases with the times the electrolyte is used, which reveals that the degradation of the electrolyte may result in a compact and passivation PPP film.     

对二氯苄-聚乙烯咪唑型聚合离子液体的制备及对煤焦油中苯酚的吸附性能

以1-乙烯咪唑和1,4-对二氯苄为原料, 通过自由基聚合和季铵化交联反应, 制得了一种新型聚合离子液体吸附剂——聚乙烯基苄基咪唑氯(P[VBnim]Cl), 并用元素分析、 凝胶渗透色谱、 红外光谱、 扫描电子显微镜、 物理吸附分析仪及热重分析等方法对其组成、 结构、 颗粒形貌、 比表面积、 孔结构及热稳定性进行了表征. 结果表明, P[VBnim]Cl 结构中对二氯苄与乙烯咪唑的摩尔比约为1∶3; 产物为米黄色蓬松粉末, 易吸水, 其初始颗粒直径为50~80 nm, 比表面积为13.86 m²/g, 平均孔径9.94 nm, 属于介孔结构材料, 初始热分解温度为274 ℃, 具有较好的热稳定性. 同时, P[VBnim]Cl对中低温煤焦油模型油中的苯酚具有优异的吸附性能, 其吸附能力是活性炭的2~10倍, 该吸附剂用乙酸乙酯再生后, 吸附效果仍然较好, 可以循环使用.     

A Simple and Efficient Electrochemical Sensor for Electrocatalytic Reduction of Nitrite Based on Poly(4‐aminoacetanilide) Film Using Carbon Paste Electrode

In this study, the electrochemical reduction of nitrite was investigated on poly(4‐aminoacetanilide) (PPAA) forming by cyclic voltammetry at the surface of carbon paste electrode. The electrochemical properties of the modified electrode have been studied by cyclic voltammetry and double potential step chronoamperometry. Results showed that in the optimum condition (pH = 0.00) the reduction of nitrite occurred at a potential about 667 mV more positive than that unmodified carbon paste electrode. This amount of electrocatalytic ability is high compared with other electrocatalysts. Using a chronoamperometric method, the catalytic rate constant (k) was calculated 8.4 × 10⁴ cm³ mol^‐1 s^‐1. Also, the electrocatalytic reduction peak currents was found to be linear with the nitrite concentration in the ranges of 5 × 10^‐4 M to 2.5 × 10^‐2 M and 2 × 10^‐5 M to 7 × 10^‐3 M with detection limits (2σ) were determined as 4.5 × 10^‐4 M and 1 × 10^‐5 M by cyclic voltammetry (CV) and hydrodynamic amperometry methods respectively. Recovery experiments exhibit the satisfactory results.