基于SiO_(2)@Fe_(3)O_(4)的磁性纳米材料分离富集谷物中痕量黄曲霉毒素B_(1)北大核心CSCD

将SiO_(2)包覆的Fe_(3)O_(4)磁性纳米材料(SiO_(2)@Fe_(3)O_(4))表面偶联识别黄曲霉毒素B_(1)(AFB_(1))的抗体(Ab),用于特异性分离富集谷物中的AFB_(1),进而与高效液相色谱-串联质谱法(HPLC-MS/MS)结合,用于大米、玉米和小麦中AFB_(1)的高效准确检测。采用微波辅助水热合成法制备得到Fe_(3)O_(4)磁性纳米颗粒,并用100μL正硅酸乙酯(TEOS)对其进行SiO_(2)的包覆,得到SiO_(2)@Fe_(3)O_(4)磁性纳米材料,随后进行抗体的偶联得到Ab-SiO_(2)@Fe_(3)O_(4);以pH=7.4的磷酸盐缓冲液(PBS)作为富集缓冲液,加入8 mg Ab-SiO_(2)@Fe_(3)O_(4),在37℃下反应10 min进行AFB_(1)的分离富集,随后采用甘氨酸-盐酸(Gly-HCl)缓冲液对Ab-SiO_(2)@Fe_(3)O_(4)分离富集的AFB_(1)进行洗涤,将洗涤液氮吹后复溶,采用高效液相色谱-串联质谱法检测。在最佳条件下,方法检测AFB_(1)的线性范围为2~50μg/L,相关系数(R^(2))>0.99,检出限为0.04μg/kg,定量限为0.13μg/kg。在4个不同加标水平下,AFB_(1)在3种谷物基质中的加标回收率为76.21%~92.85%,RSD≤5.29%。大米、玉米和小麦等实际谷物样品中AFB_(1)的测定结果显示,在1个小麦样品和2个玉米样品中检出AFB1,其含量分别为0.38、0.13和0.47μg/kg,其他样品中并未发现AFB_(1)。方法将磁性纳米材料与HPLC-MS/MS相结合,实现了AFB_(1)的高效分离富集,富集材料成本低廉,储存性能好,在30 min内即可完成前处理过程,可在较短的时间内实现大批量样品的实际分析,在谷物中真菌毒素的检测方面具有良好的应用前景。     

基于磁珠-适配体的高效液相色谱-串联质谱法检测食品中的黄曲霉毒素B1

本文构建了特异性识别黄曲霉毒素B₁(AFB₁)的磁珠-适配体,并与高效液相色谱-串联质谱联用(LC-MS/MS),建立食品中AFB₁的定量检测方法。 利用碳二亚胺盐酸盐(EDC)活化法,将羧基磁珠进行活化。 活化后的羧基磁珠与5'端氨基修饰的适配体进行孵育结合,通过酰胺反应将适配体共价连接在羧基磁珠表面,固定在磁珠表面的适配体作为捕捉探针将样品提取液中的AFB₁分离,通过LC-MS/MS对AFB₁进行定性和定量分析。 检测结果表明:AFB₁在浓度0.25～25 ng/mL呈良好的线性关系,相关系数R²=0.999,定量检出限为0.25 ng/mL,回收率达到80.3%～92.5%,相对标准偏差(RSD)低于8%。 该方法操作简单、快速便捷、可痕量地检测AFB₁,所制备的磁珠-适配体可重复利用,为定量检测AFB₁提供了另一种技术支持。     

免疫磁珠富集净化-超高效液相色谱法同时测定陈皮中4种黄曲霉毒素

将ProtElut NHS(N-羟基丁二酰亚胺)偶联磁珠与抗黄曲霉毒素总量单克隆抗体偶联得到黄曲霉毒素总量免疫磁珠,其具有较好的分散性,良好的磁性能和特异的选择性。本文建立了陈皮中4种黄曲霉毒素的免疫磁珠富集净化,超高效液相色谱(UPLC)检测方法。样品经甲醇-PBS缓冲溶液(2:8, v/v)提取后用免疫磁珠富集净化,1 mL甲醇洗脱;经ACQUITY UPLC HSS T3 C₁₈色谱柱(100 mm×2.1 mm, 1.8 μm)分离,以水和甲醇为流动相梯度洗脱,采用汞/氙灯荧光检测器测定。实验结果表明,4种黄曲霉毒素在各自的线性范围内峰面积与其质量浓度线性关系良好,相关系数(r²)大于0.999;检出限(信噪比为3)为0.013~0.038 μg/kg,定量限(信噪比为10)为0.044~1.2 μg/kg;平均回收率为63.9%~115.0%,相对标准偏差为0.4%~14.2%,均符合痕量分析的要求。该方法简单快速、准确可靠,可用于陈皮中4种黄曲霉毒素的测定。     

Chronoamperometry Measurement for Rapid Cucumber Mosaic Virus Detection in Plants

Cucumber mosaic virus (CMV) causes major losses to agricultural and horticultural crops around the world. Hence, a rapid assay for the detection of CMV which can be employed in both laboratory and field is essential. A portable electrochemical immunosensor system for the detection of CMV, based on immobilized CMV specific antibodies conjugated with gold nanoparticle was developed for this purpose. The conjugated antibodies were added with polymer and deposited onto carbon screen printed working electrodes. Optimization of the modified surface immunosensor was performed using sandwich immunoassay format (ELISA). The initial ELISA result for the standard curve development showed a limit of detection down to 0.1mg/mL. Subsequently, the immunosensor was tested for cross reactivity with other plant pathogens. The performance of the electrochemical immunosensor revealed that it has a high selectivity in sample matrix with other organism. This immunosensor provides a promising technology for simple and sensitive detection system that is essential in rapid detection of plant pathogens.     

Flow-injection immuno-bioassay for interleukin-6 in humans based on gold nanoparticles modified screen-printed graphite electrodes

A flow-injection electrochemical immunoassay system based on a disposable immunosensor for the determination of interleukin-6 (IL-6) was proposed. The immunosensor was prepared by entrapping horseradish peroxidase (HRP)-labeled IL-6 antibody into gold nanoparticles-modified composite membrane at a screen-printed graphite electrode. With a non-competitive immunoassay format, the immunosensor was inserted in the flow system with an injection of sample, and the injected sample containing IL-6 antigen was produced transparent immunoaffinity reaction with the immobilized HRP-labeled IL-6 antibody. The formed antigen–antibody complex inhibited partly the active center of HRP, and decreased the immobilized HRP to H₂O₂ reduction. The performance and factors influencing the performance of the immunosensor were investigated. Under optimal conditions, the current change obtained from the labeled HRP relative to thionine–H₂O₂ system was proportional to the IL-6 concentration in the range of 5–100 ng L⁻¹ with a detection limit of 1.0 ng L⁻¹ (at 3δ). The flow-injection immunoassay system could automatically control the incubation, washing and measurement steps with acceptable reproducibility and good stability. Moreover, the proposed immunosensors were used to analyze IL-6 in human serum specimens. Analytical results of clinical samples show the developed immunoassay has a promising alternative approach for detecting IL-6 in the clinical diagnosis.     

黄曲霉素B_2分子吸附在银团簇的表面增强拉曼散射机理

采用密度泛函理论(DFT)B3LYP方法,6-311G(d,p)(C,H,O)/LANL2DZ(Ag)基组,计算了黄曲霉素B2(AFB2)分子吸附在Ag2团簇的表面增强拉曼散射(SERS)光谱和预共振拉曼光谱,并与实验结果比较.结果显示:AFB2分子在基态Ag2团簇表面吸附时,增强因子最大达到102,对应吡喃(pyrane)环C=O伸缩振动,主要是由AFB2分子周围化学环境改变而引起的基态静极化率改变导致的化学增强.不同激发波长下的AFB2分子预共振拉曼光谱的增强强度不同:电荷转移态激发波长为1144和544 nm时拉曼信号增强了102倍,而选择电荷转移预共振波长432和410 nm作为入射光时,其拉曼信号增强了104倍,增强机理为银团簇和黄曲霉素分子之间的电荷转移共振增强.因此通过改变入射光波长,选择电荷转移共振激发波长,更有利于强致癌物AFB2分子的痕量检测.     

Evaluation of electrochemical hydride generation for spectrophotometric determination of As(III) by silver diethyldithiocarbamate

A batch electrochemical hydride generation system was developed for the spectrophotometric determination of inorganic As(III) by silver diethyldithiocarbamate. This method is based on electrochemical reduction of As to Arsine (AsH₃) in acidic media and on the subsequent reaction of AsH₃ with silver diethyldithiocarbamate to give an absorbing complex at 525 nm. The electrochemical generator consisted of a cathode cell separated from the anode cell by a porous glass frit and was operated with a constant direct current. A pre-activated graphite rod was used as cathode material for the production of AsH₃. The parameters related to the electrochemical hydride generation were investigated. Under optimized conditions, only As(III) can be converted to AsH₃ and a pre-reducing stage is required for total inorganic As analysis. The characteristic data of the electrochemical hydride generation and common hydride generation by NaBH₄ were compared. Also, the effects of interferences and their elimination were investigated. An absolute detection limit of 0.5 μg (3s_b) and a concentration detection limit of 0.05 μg/ml were obtained using a 10 ml sample volume. The relative standard deviation for five replicate analysis of 30 μg As(III) was 1.2%. The accuracy and recovery of the method were demonstrated by analysing spiked artificial sea water and tap water.     

Synthesis and electrochemical properties of Zn3V3O8 as novel anode material

Zn₃V₃O₈ two-dimensional micro sheets are successfully synthesized by combination of solvothermal method and heat treatment. The Zn₃V₃O₈ has better electrochemical performances after calcinations.     

Transition metal binding properties of the redox-active 1,4,7,10,13,16-hexa(ferrocenylmethyl)-1,4,7,10,13,16-hexaazacyclooctadecane and its electrochemical behaviour in a non-aqueous solvent

Solution studies to elucidate the coordination behaviour and the electrochemical response of the ferrocene-functionalized polyazamacrocycle 1,4,7,10,13,16-hexa(ferrocenylmethyl)-1,4,7,10,13,16-hexaazacyclooctadecane (L¹) by potentiometric methods and electrochemical techniques have been carried out. Potentiometric methods in the presence of Cd²⁺, Hg²⁺, Pb²⁺ and Zn²⁺ were carried out in 1,4-dioxane/water (70:30 v/v, 25°C, 0.1 mol dm⁻³ KNO₃). Electrochemical studies were carried out in acetonitrile/dichloromethane (50:50 v/v, 25°C, 0.1 mol dm⁻³ TBAClO₄) in the presence of transition metal ions and anions.     

基于花状铂纳米颗粒构建的电致化学发光免疫传感器用于检测载脂蛋白A1

采用一锅合成法制备了新型的具有大比表面积的花状铂纳米颗粒（PtNFs）,并构建了一个高灵敏电致化学发光（ECL）免疫传感器用于检测载脂蛋白A1（Apo-A1）. 该PtNFs用于吸附二抗（anti-Apo-A1）,并用葡糖糖氧化酶（GOD）封闭其表面的非特异性位点,最终制备了PtNFs@anti-Apo-A1@GOD信号探针. 当Apo-A1存在时,通过夹心免疫反应将制备的信号探针捕获于电极表面,并将所制得的电极置于含有葡萄糖的过硫酸根底液中检测. GOD催化葡萄糖产生H₂O₂,H₂O₂在PtNFs的催化下分解并在电极表面原位产生O₂,所产生的O₂能够催化过硫酸根-氧气体系的电致化学发光反应,放大发光信号,提高检测灵敏度. 该传感器在0.1ng•mL^-1 ~ 100 ng•mL^-1范围内对Apo-A1有良好的线性响应,检测下限达到0.03ng•mL^-1,有望应用于临床分析诊断.     

Electrochemical Immunoassay of E. coli in Urban Sludge Using Electron Mediator-mediated Enzymatic Catalysis and Gold Nanoparticles for Signal Amplification

An electrochemical immunosensor was developed for sensitive assay of E. coli in urban sludge, in which electron mediator-mediated enzymatic catalysis and gold nanoparticles(AuNPs) were utilized for signal amplification. The immnuosensing platform chitosan-thionine(chit-thio)/poly(amidoamine) dendrimer-encapsulated AuNPs [PAMAM(Au)] composites were first prepared from chit-thio and PAMAM(Au) using the layer-by-layer method to provide a matrix for high-stability and high-bioactivity bindings of the capture antibody(_cAb). Moreover, the {_dAb-AuNPs-HRP} nanoprobes were designed to exploit the amplification effect of the carrier AuNPs due to the loading amounts of horseradish peroxidase(HRP) and the detection antibody(_dAb). The sandwich-type immunoassay was then successfully used to assay E. coli based on the oxidation of thionine as a result of H₂O₂-induced enzymatic catalytic reaction by HRP. This study presents a powerful tool in electrochemical immunoassay for E. coli detection with rapid response, high-sensitivity and high-specificity, providing a potential new tool for feasibility assessment of sludge recycle.     

Sensitivity improvement of a sandwich-type ELISA immunosensor for the detection of different prostate-specific antigen isoforms in human serum using electrochemical impedance spectroscopy and an ordered and hierarchically organized interfacial supramolecular architecture

A gold millielectrode (GME) functionalized with a mixed (16-MHA + EG₃SH) self-assembled monolayer (SAM) was used to fabricate an indirect enzyme-linked immunosorbent assay (ELISA) immunosensor for the sensitive detection of prostate-specific antigen (PSA), a prostate cancer (PCa) biomarker, in human serum samples. To address and minimize the issue of non-specific protein adsorption, an organic matrix (amine-PEG₃-biotin/avidin) was assembled on the previously functionalized electrode surface to build up an ordered and hierarchically organized interfacial supramolecular architecture: Au/16-MHA/EG₃SH/amine-PEG₃-biotin/avidin. The electrode was then exposed to serum samples at different concentrations of a sandwich-type immunocomplex molecule (^BtnAb-Ag_PSA-^HRPAb), and its interfacial properties were characterized using electrochemical impedance spectroscopy (EIS). Calibration curves for polarization resistance (R_P) and capacitance (1/C) vs. total and free PSA concentrations were obtained and their analytical quality parameters were determined. This approach was compared with results obtained from a commercially available ELISA immunosensor. The results obtained in this work showed that the proposed immunosensor can be successfully applied to analyze serum samples of patients representative of the Mexican population.     

Ag/C包覆对Li[Li0.2Mn0.54Ni0.13Co0.13]O2电化学性能的影响

运用共沉淀和元素化学沉积相结合的方法,制备出了具有Ag/C包覆层的层状富锂固溶体材料Li[Li0.2Mn0.54Ni0.13Co0.13]O2.通过X射线衍射(XRD)、场发射扫描电子显微镜(SEM)、透射电子显微镜(TEM)、恒流充放电、循环伏安(CV),电化学阻抗谱(EIS)和X射线能量散射谱(EDS)方法,研究了Ag/C包覆层对Li[Li0.2Mn0.54Ni0.13Co0.13]O2电化学性能的影响.结果表明,Ag/C包覆层的厚度约为25 nm,Ag/C包覆在保持了固溶体材料α-NaFeO2六方层状晶体结构的前提下,显著地改善了Li[Li0.2Mn0.54Ni0.13Co0.13]O2的电化学性能.在2.0-4.8 V(vs Li/Li+)的电压范围内,首次放电(0.05C)容量由242.6 mAh·g-1提高到272.4 mAh·g-1,库仑效率由67.6%升高到77.4%;在0.2C倍率下,30次循环后,Ag/C包覆的电极材料容量为222.6 mAh·g-1,比未包覆电极材料的容量高出14.45%;包覆后的电极材料在1C下的容量仍为0.05C下的81.3%.循环伏安及电化学交流阻抗谱研究表明,Ag/C包覆层抑制了材料在充放电过程中氧的损失,有效降低了Li[Li0.2Mn0.54Ni0.13Co0.13]O2颗粒的界面膜电阻与电化学反应电阻.     

Comparison of Electrochemical and Spectro-Photometric Detection in Hrp-Based Elisa for Tobacco Mosaic Virus

ABSTRACT

An electrochemical method was compared with a spectrophotometric method using a horseradish peroxidase (HRP)-based indirect enzyme-linked immunosorbent assay (ELISA) with direct antigen coating (DAC) technique for the determination of tobacco mosaic virus (TMV). In substrates for a spectrophotometric HRP-based ELISA method, was selected as the substrate for the electrochemical method as well as the spectrophotometric method. 2, 3-Diaminophenazine is the product of the oxidation of OPD with H₂O₂ catalyzed by HRP in the selected conditions. It is electroactive and has a sensitive voltammetric response at -0.93 V (vs. Ag/AgCl) in alkaline solution. The detection limit of free HRP, by second-order derivative linear-sweep voltammetry, is 1.1 mU/L. This method can be further used to detect TMV antigen. The peak current is linear with TMV concentration in the range of 0.25-5.0 ng/mL. The detection limit for TMV is 0.25 ng/mL and the highest dilution ratio detected for the infected leaf sap is 1:102400. Compared with the classical OPD ELISA spectrophotometric method, the detection limits of the electrochemical method for the clarified TMV and the infected leaf sap detection are about ten and four times lower, respectively.     

Nanosilver and DNA-functionalized immunosensing probes for electrochemical immunoassay of alpha-fetoprotein

A simple and sensitive electrochemical immunosensor for a one-step immunoassay for alpha-fetoprotein (AFP) was designed using silver nanoparticles and double-stranded DNA as matrices. The detection was based on the change in the electron transfer resistance before and after the antigen-antibody reaction by using electrochemical impedance spectroscopy. Under optimal conditions, the resistance shift of the immunosensor is proportional to the AFP concentration in the range 3.5 –360 ng·mL^?1 with a detection limit of 1.5 ng·mL^?1 (at 3σ). The immunosensor exhibits high sensitivity, good reproducibility and stability. Results obtained for clinical serum samples by the immunosensor are in accordance with those determined by spectrophotometric enzyme-linked immunosorbent assays.     

A competitive microcystin-LR immunosensor based on Au NPs@metal-organic framework (MIL-101)

A porous metal organic frameworks (MOFs) material (MIL-101) based on trivalent chromium skeleton were synthesized by hydrothermal synthesis method, and loaded with Au nanoparticles (Au NPs) to prepare Au NPs@MIL-101 composite materials which were used as a marker to label anti microcystin-LR (Anti-MC-LR). The composite materials have strong catalytic properties to the oxidation of ascorbic acid. Anti-MC-LR was immobilized on glassy carbon electrode surface using electrodeposition graphene oxide (GO) as a fixed matrix to construct a competitive microcystin-LR immunosensor.     

Biomolecule‐Doped Organic/Inorganic Hybrid Nanocomposite Film for Label‐Free Electrochemical Immunoassay of α‐1‐Fetoprotein

A new electrochemical immunosensor for the detection of α‐1‐fetoprotien (AFP) was developed based on AFP antibody (anti‐AFP)‐functionalized organic/inorganic hybrid nanocomposite membrane. To fabricate such a hybrid composite membrane, 3,4,9,10‐perylenetetracarboxylic acid‐bound thionine molecules (PTCTH) were initially doped into titania colloids (TiO₂), and then gold nanoparticles and anti‐AFP were immobilized onto the composite film in turn. Comparison with the electrode fabricated only with thionine not 3,4,9,10‐perylenetetracarboxylic acid, the immunosensor with PTCTH exhibited high sensitivity and fast electron transfer. The presence of gold nanoparticles provided a good microenvironment for the immobilization of biomolecules, enhanced the surface coverage of protein, and improved the sensitivity of the immunosensor. The modified process was characterized by scanning electron microscope (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The surface topography of the membrane was investigated by scanning electron microscopy (SEM). Under optimal conditions, the proposed immunosensor exhibited a wide linear range from 2.5 to 200.0 ng/mL towards AFP with a detection limit of 0.5 ng/mL (S/N=3). The stability, reproducibility and precision of the immunosensor were acceptable. Comparison with the conventional enzyme‐linked immunosorbent assay (ELISA), the present method did not require more labeled procedures and washing steps. Significantly, the detection methodology provides a promising approach for other proteins or biosecurities.     

Preparation of TiO2 Nanoflakes and Their Influence on Lithium Ion Battery Storage Performance

TiO₂ nanoflakes were prepared by hydrothermal precipitation method using Ti(SO₄)₂ as titanium source and NaOH solution as alkaline medium. Their surface morphology, grain size measured after high temperature calcination and effect on the electrochemical performance of Li ion battery were discussed. TiO₂ nanoflakes were characterizated by means of transmission electron microscopy(TEM), X^-ray powder diffraction(XRD), N₂ adsorption-desorption isothermal assay, cyclic voltammetry(CV) and cycle performance test. The result of electrochemical performance test shows that the prepared TiO₂ nanoflakes have high discharge specific capacity and good cycle performance. Discharge specific capacity for the first circle at the discharge rate of 0.1 C is 261.5 mA·h·g^-1. After 90 cycles, the discharge capacity reduces to 172.2 mA·h·g^-1.     

HPLC determination of Vitamin D(3) and its metabolite in human plasma with on-line sample cleanup

A HPLC method with automated column switching and UV-diode array detection is described for the simultaneous determination of Vitamin D₃ and 25-hydroxyvitamin D₃ (25-OH-D₃) in a sample of human plasma. The system uses a BioTrap precolumn for the on-line sample cleanup. A sample of 1 ml of human plasma was treated with 2 ml of a mixture of ethanol–acetonitrile (2:1 (v/v)). Following centrifugation, the supernatant was evaporated to dryness under a stream of dry and pure nitrogen. The residue was reconstituted in 250 μL of a solution of methanol 5 mmol l⁻¹ phosphate buffer, pH 6.5 (4:1 (v/v)), and a 200 μl aliquot of this solution was injected onto the BioTrap precolumn. After washing during 5 min with a mobile phase constituted by a solution of 6% acetonitrile in 5 mmol l⁻¹ phosphate buffer, pH 6.5 (extraction mobile phase), the retained analytes were then transferred to the analytical column in the backflush mode. The analytical separation was then performed by reverse-phase chromatography in the gradient elution mode with the solvents A and B (Solvent A: acetonitrile–phosphate buffer 5 mmol l⁻¹, pH 6.5; 20:80 (v/v); solvent B: methanol–acetonitrile–tetrahydrofuran, 65:20:15 (v/v)). The compounds of interest were detected at 265 nm. The method was linear in the range 3.0–32.0 ng ml⁻¹ with a limit of quantification of 3.0 ng ml⁻¹. Quantitative recoveries from spiked plasma samples were between 91.0 and 98.0%. In all cases, the coefficient of variation (CV) of the intra-day and inter-day-assay precision was ≤2.80%. The proposed method permitted the simultaneous determination of Vitamin D₃ and 25-OH-D₃ in 16 min, with an adequate precision and sensitivity. However, the overlap of the sample cleanup step with the analysis increases the sampling frequency to five samples h⁻¹. The method was successfully applied for the determination of Vitamin D₃ and 25-OH-D₃ in plasma from 46 female volunteers, ranging from 50 to 94 years old. Vitamin D₃ and 25-OH-D₃ concentrations in plasma were found from 4.30–40.70 ng ml⁻¹ (19.74 ± 9.48 ng ml⁻¹) and 3.1–36.52 ng ml⁻¹ (7.13 ± 7.80 ng ml⁻¹), respectively. These results were in good agreement with data published by other authors.     

锆掺杂以提升LiNi0.5Co0.2Mn0.3O2正极材料的高温电化学性能

为解决LiNi_0.5Co_0.2Mn_0.3O₂正极材料在高温下循环性能差的问题,本文通过固相法对材料进行锆掺杂改性,研究了不同掺杂量对LiNi_0.5Co_0.2Mn_0.3O₂晶体结构和电化学性能的影响。研究表明,当锆掺杂量为1% (x)时,可以降低LiNi_0.5Co_0.2Mn_0.3O₂结构中的Li⁺/Ni²⁺离子混排,有助于材料电化学性能的提高,尤其是高温循环性能。在25 ℃、3.0-4.3 V下, Li(Ni_0.5Co_0.2Mn_0.3)_0.99Zr_0.01O₂在1C循环95次后容量保持率为92.13%,优于未掺杂样品(87.61%)。在55 ℃下, Li(Ni_0.5Co_0.2Mn_0.3)_0.99Zr_0.01O₂在1C循环115次后容量保持率仍有82.96%,远高于未掺杂样品(67.63%)。因此,少量锆掺杂对提升LiNi_0.5Co_0.2Mn_0.3O₂的高温循环性能有积极作用。