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1.
应用主客体分子识别技术构建一种无需固定DNA探针的电化学检测DNA的新方法。分别采用间甲基苯甲酸标记和金纳米颗粒标记的两段DNA探针,使用β-环糊精修饰电极对杂交结构进行识别和电化学检测。采用差分脉冲伏安法(DPV)实现对特定序列DNA的定量检测。在优化的条件下,该体系表现出良好的灵敏度和选择性,对特定序列DNA的检测限为1.8×10-13mol/L。  相似文献   

2.
功能化纳米金放大的DNA电化学传感器研究   总被引:7,自引:0,他引:7  
研究了DNA夹心杂交和直接杂交体系,将功能化纳米金引入到标记有生物素的杂交双链上,制成具有电化学活性和纳米金放大作用的DNA电化学传感器,采用循环伏安法测试.在夹心杂交体系中,靶点DNA浓度与阳极峰电流关系曲线的相对标准偏差为3.0%~13.0%,在浓度为6.9×10-3~0.14nmol/L范围内得到良好的线性关系,检测限达到2.0×10-3nmol/L,实现了对单碱基突变的高灵敏检测和序列识别.直接杂交检测限为2.5×10-4mol/L,分别在2.5×10-4~5.0×10-3nmol/L和5.0×10-3~10nmol/L范围内得到峰电量与浓度的良好线性关系.并比较这两种体系.  相似文献   

3.
功能化纳米金增强的DNA电化学检测和序列分析   总被引:6,自引:0,他引:6  
李金花  胡劲波 《化学学报》2004,62(20):2081-2088,F010
用冠以大量二茂铁的纳米金微粒 /抗生蛋白链菌素结合物为标记物 ,将其标记于生物素修饰的寡聚核苷酸片段上 ,制成了具有电化学活性和纳米金放大作用的DNA电化学生物传感器 .首先采用巯基DNA和巯基烷烃混合自组装膜制备了金修饰电极 ,将探针DNA分子固定在了电极表面 ,运用杂交原则结合靶点分子在电极表面形成了双螺旋的DNA链 ,然后借助抗生蛋白链菌素和生物素之间的强亲和作用 ,引入了功能化的纳米金 .通过伏安法测定了修饰在纳米金上的二茂铁的氧化还原电流 ,可以识别和测定溶液中互补的靶点DNA ,17 mer靶点DNA的浓度在 0 .0 0 1~ 10nmol/L范围内有线性关系 ,检测限可达 0 .75× 10 -12 mol/L .  相似文献   

4.
CdTe量子点标记的DNA电化学传感器的研究   总被引:2,自引:1,他引:1  
利用碳纳米管和CdTe量子点(QDs)组装的电化学传感器,建立了一种识别DNA的新方法.将氨基修饰的单链DNA探针共价键合固定在带有羧基的碳纳米管修饰的金电极上,然后与CdTe QDs标记的目标DNA进行杂交.利用差分脉冲法(DPV)和循环伏安法对目标DNA的固定和杂交进行表征,通过电活性指示剂柔红霉素(DNR)的DPV峰电流变化,对互补DNA、非互补DNA和单碱基错配DNA序列进行识别.与未标记CdTe QDs的目标DNA相比,标记CdTe QDs的目标DNA序列的电流响应灵敏度明显提高.DNA电化学传感器检测的优化条件:DNR的浓度为1.67×10-5 mol/L,DNA杂交时间为80 min,杂交温度为55 ℃.在1.0×10-13 ~1.0×10-8 mol/L范围,目标DNA浓度的对数值与其响应的DPV信号(还原峰电流)呈线性关系,检出限为3.52×10-14 mol/L(S/N=3,n=9),线性方程为ΔI=50.22+3.567 lgcDNA,相关系数为0.996 6.对1.0×10-10 mol/L的目标DNA样品进行重复测定,相对标准偏差为4.8%(n=5),重复性良好.  相似文献   

5.
用模板法在氧化铟锡(ITO)电极上制备具有三维有序多孔结构的金掺杂纳米二氧化钛修饰电极(3DOM GTD/ITO),扫描电镜(SEM)结果表明,制备的修饰电极三维结构规整有序、孔径均一。将标记有二茂铁(Fc)的DNA探针修饰到3DOM GTD/ITO电极上构建了一种新的标记型DNA生物传感器,通过Fc在DNA探针杂交前后的电化学信号变化可识别目标靶序列。采用循环伏安(CV)、示差脉冲(DPV)和交流阻抗(EIS)等方法对DNA探针在电极表面的固定和杂交进行表征。实验结果表明,该DNA生物传感器可以成功地识别乳腺癌基因靶序列,Fc的氧化还原电流与靶序列浓度在8.0×10-7~1.0×10-5 mol/L范围内呈线性关系,线性相关系数为0.9908,检测限为5.2×10-7 mol/L。  相似文献   

6.
基于金纳米颗粒(AuNPs)比表面积大、 尺寸小和能够承载大量DNA片段的特点, 建立了一种免标记、 简便、 快速检测DNA聚合酶Klenow fragment exo-(KF-)的电化学方法. 首先将巯基化的DNA引物片段修饰在金电极上, 然后加入模板DNA链以及修饰有报告DNA链的金纳米颗粒(AuNPs-DNA), 模板DNA链能同时与DNA引物片段和修饰在AuNPs上的报告DNA链进行互补杂交形成"三明治"结构, 从而将AuNPs-DNA修饰在电极表面; 当加入电活性物质钌铵(RuHex)后, RuHex可通过静电吸附作用结合在DNA上. AuNPs上修饰的报告DNA链能够吸附大量RuHex, 导致电化学信号放大. 当加入脱氧核糖核苷三磷酸(dNTPs)以及KF-聚合酶后, 引物片段发生延伸反应, 将与模板DNA链杂交的AuNPs-DNA竞争下来, 带走大量的RuHex, 使电信号降低, 从而实现对聚合酶的检测. 实验结果表明, 利用该方法可以检测到5 U/mL的KF-.  相似文献   

7.
基于硫化镉纳米团簇标记DNA电化学传感的研究   总被引:5,自引:2,他引:3  
祝宁宁  张爱平  何品刚  方禹之 《化学学报》2003,61(10):1682-1685
合成了表面具有自由羧基的硫化镉纳米团簇,以乙基-(3-二甲基丙基)碳二 亚胺盐酸盐为偶联活化剂,将其标记于人工合成的5'端氨基修饰的寡聚核苷酸片段 上,制备成CdS纳米团簇标记DNA探针,该寡聚核苷酸片段与大肠杆菌肠毒素基因相 关。在一定的条件下,使基与固定晨玻碳电极表面的待测DNA序列进行杂交反应, 利用阳极溶出示差脉冲伏安法(ASDPV)间接测定Cd的量,实现对互补、非互补 DNA片段的识别和电化学检测,从而对大肠杆菌肠毒素基因片段识别和检测。  相似文献   

8.
基于发夹型核酸探针的高特异性识别能力以及电活性物质与DNA磷酸骨架间的静电作用,以发夹型核酸作为分子识别探针,电活性物质六氨合钌(RuHex)作为杂交指示剂,构建了一种非标记型检测p53抑癌基因的电化学DNA生物传感器.实验结果表明,在10 μmol/L RuHex溶液中,该传感器对目标DNA具有灵敏的电化学响应,电化...  相似文献   

9.
本文以人参ITS及518 s基因上的SNP位点为检测对象,利用分子识别作用构建了一种电化学传感器,成功地对人参、西洋参进行了品种鉴别。本文设计合成了一种双标记DNA探针(DLP),该探针的一端标记了4-4-二甲氨基苯基偶氮苯甲酸(dabcyl)作为客体分子,另一端标记了金纳米颗粒作为电化学杂交指示剂。同时使用α-CD/MCNTs/GCE电极作为工作电极。由于DLP的茎环结构,只有在DLP与目标DNA杂交后,DLP上的dabcyl分子进入修饰电极表面的α-CD空腔中,进而DLP被α-CD修饰电极捕获。并且,通过金纳米颗粒的AuCl4-的电化学还原电流信号。可灵敏检测4.6×10-10mol.L-1的目标DNA。  相似文献   

10.
韩苗苗  王萍  席守民 《分析测试学报》2020,39(12):1466-1472
该文以DNA四面体纳米结构探针(TSP)为捕获探针,将辣根过氧化物酶标记的IgG抗体结合在纳米金颗粒表面(AuNPs-IgG-HRP)作为信号分子,构建了一种新型DNA甲基化电化学传感器。利用一步热变性法组装成TSP后,通过Au—S键固定在修饰纳米金颗粒的金电极表面,经过靶标DNA杂交、5-甲基胞嘧啶(5-mc)抗体及AuNPs-IgG-HRP结合后,用差分脉冲伏安法(DPV)进行检测。采用循环伏安法(CV)和电化学阻抗谱(EIS)对修饰电极的构建过程进行电化学表征。探究了杂交时间、5-mc抗体浓度、IgG-HRP加入体积、氢醌(HQ)和过氧化氢(H2O2)浓度对传感器的影响。在最佳条件下,该传感器对甲基化DNA的线性响应范围为1.0×10-15~1.0×10-10 mol/L,检出限(S/N=3)为4.4×10-16 mol/L。该传感器具有良好的选择性和稳定性,为DNA甲基化检测提供了新方法。  相似文献   

11.
Sun Z  Qiang W  Li H  Hao N  Xu D  Chen HY 《The Analyst》2011,136(3):540-544
In this work a novel microdevice sensor has been developed by plating gold on the PDMS surface to generate a sandwich-type gap electrode for DNA detection. The microdevice utilizes a gold band electrode-PDMS-gold band electrode configuration and the minimum detectable volume could be as low as 5 μL. The 20 μm PDMS-based gap was chemically modified with DNA capture probes and DNA sandwich hybrids were formed with the addition of DNA target and silver nanoparticle probes. To increase detection sensitivity, parallel detection zones have been developed in which the relevant resistances decrease substantially upon hybridyzation. By measuring the change in electrical conductivity, the DNA target in the concentration range of 1000-0.1 nM can be assayed and the limit of lowest detectable concentration was achieved at 0.01 nM.  相似文献   

12.
A sensitive electrochemical method for the detection of DNA hybridization based on the probe labeled with multiwall carbon‐nanotubes (MWNTs) loaded with silver nanoparticles (Ag‐MWNTs) has been developed. MWNTs were electroless‐plated with a large number of silver nanoparticles to form Ag‐MWNTs. Probe single strand DNA (ss‐DNA) with a thiol group at the 3′‐terminal labeled with Ag‐MWNTs by self‐assembled monolayer (SAM) technique was employed as an electrochemical probe. Target ss‐DNA with a thiol group was immobilized on a gold electrode by SAM technique and then hybridized with the electrochemical probe. Binding events were monitored by differential pulse voltammetric (DPV) signal of silver nanoparticles. The signal difference permitted to distinguish the match of two perfectly complementary DNA strands from the near perfect match where just three base pairs were mismatched. There was a linear relation between the peak current at +120 mV (vs. SCE) and complementary target ss‐DNA concentration over the range from 3.1×10?14 to 1.0×10?11 mol/L with a detection limit of 10 fmol/L of complementary target ss‐DNA. The proposed method has been successfully applied to detection of the DNA sequence related to cystic fibrosis. This work demonstrated that the MWNTs loaded with silver nanoparticles offers a great promising approach for sensitive detection of DNA hybridization.  相似文献   

13.
We use colloidal Au to enhance the DNA immobilization amount on a gold electrode and ultimately lower the detection limit of our electrochemical DNA biosensor. Self-assembly of approximately 16-nm diameter colloidal Au onto a cysteamine modified gold electrode resulted in an easier attachment of an oligonucleotide with a mercaptohexyl group at the 5′-phosphate end, and therefore an increased capacity for nucleic acid detection. Quantitative results showed that the surface densities of oligonucleotides on the Au colloid modified gold electrode were approximately (1–4)×1014 molecules cm−2. Hybridization was induced by exposure of the ssDNA-containing gold electrode to ferrocenecarboxaldehyde labeled complementary ssDNA in solution. The detection limit is 5×10−10 mol l−1 of complementary ssDNA, which is much lower than our previous electrochemical DNA biosensors. The Au nanoparticle films on the Au electrode provide a novel means for ssDNA immobilization and sequence-specific DNA detection.  相似文献   

14.
The unique binding event between Escherichia coli single-stranded DNA binding protein (SSB) and single-stranded oligonucleotides conjugated to gold (Au) nanoparticles is utilized for the electrochemical detection of DNA hybridization. SSB was attached onto a self-assembled monolayer (SAM) of single-stranded oligonucleotide modified Au nanoparticle, and the resulting Au-tagged SSB was used as the hybridization label. Changes in the Au oxidation signal was monitored upon binding of Au tagged SSB to probe and hybrid on the electrode surface. The amplified oxidation signal of Au nanoparticles provided a detection limit of 2.17 pM target DNA, which can be applied to genetic diagnosis applications. This work presented here has important implications with regard to combining a biological binding event between a protein and DNA with a solid transducer and metal nanoparticles.  相似文献   

15.
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.  相似文献   

16.
基于纳米金探针和基因芯片的DNA检测新方法   总被引:2,自引:0,他引:2  
包华  贾春平  周忠良  金庆辉  赵建龙 《化学学报》2009,67(18):2144-2148
运用荧光纳米金探针和基因芯片杂交建立一种新的DNA检测方法. 荧光纳米金探针表面标记有两种DNA探针: 一种为带有Cy5荧光分子的信号探针BP1, 起信号放大作用; 另一种为与靶DNA一部分互补的检测探针P532, 两种探针比例为5∶1. 当靶DNA存在时, 芯片上捕捉探针(与靶DNA的另一部分互补)通过碱基互补配对结合靶DNA, 将靶DNA固定于芯片上; 荧光纳米金探针通过检测探针与靶DNA及芯片结合, 在芯片上形成“三明治”复合结构, 最后通过检测信号探针上荧光分子的信号强度来确定靶DNA的量. 新方法检测灵敏度高, 可以检测浓度为1 pmol/L的靶DNA, 操作简单, 检测时间短. 通过改进纳米金探针的标记和优化杂交条件, 可进一步提高核酸检测的灵敏度, 这将在核酸检测方面具有重要的应用价值.  相似文献   

17.
《Electroanalysis》2004,16(19):1628-1631
In this work, we report a simple, rapid and sensitive approach for the electrochemical gold nanoparticle‐based DNA detection with an electrocatalytic silver deposition process. The catalytic and preferential silver electrodeposition on gold nanoparticle surfaces using an indium tin oxide (ITO) electrode at certain potentials, without any chemical pretreatments of the electrode, is demonstrated. More importantly, the application of this methodology for hybridization transduction is explored. The ITO electrode surface is first coated with an electroconductive polymer, poly(2‐aminobenzoic acid), to enable the chemical attachment of avidin molecules for the subsequent probe immobilization. The hybridization of the target with the probe in turn permits the binding of the gold nanoparticle labels to the transducer surface via biotin‐streptavidin interaction. The amount of bound gold labels, which is proportional to the amount of the target, is determined by the electrocatalytic silver deposition process. A significant improvement of the signal‐to‐background ratio is achieved with this scheme compared to the conventional chemical hydroquinone‐based silver deposition process.  相似文献   

18.
This study demonstrates a highly sensitive sensing scheme for the detection of low concentrations of DNA, in principle down to the single biomolecule level. The previously developed technique of electrochemical current amplification for detection of single nanoparticle (NP) collisions at an ultramicroelectrode (UME) has been employed to determine DNA. The Pt NP/Au UME/hydrazine oxidation reaction was employed, and individual NP collision events were monitored. The Pt NP was modified with a 20-base oligonucleotide with a C6 spacer thiol (detection probe), and the Au UME was modified with a 16-base oligonucleotide with a C6 spacer thiol (capture probe). The presence of a target oligonucleotide (31 base) that hybridized with both capture and detection probes brought a Pt NP on the electrode surface, where the resulting electrochemical oxidation of hydrazine resulted in a current response.  相似文献   

19.
Fan A  Lau C  Lu J 《The Analyst》2008,133(2):219-225
A sensitive chemiluminescent (CL) detection of sequence-specific DNA has been developed by taking advantage of a magnetic separation/mixing process and the amplification feature of colloidal gold labels. In this protocol, the target oligonucleotides are hybridized with magnetic bead-linked capture probes, followed by the hybridization of the biotin-terminated amplifying DNA probes and the binding of streptavidin-coated gold nanoparticles; the nanometer-sized gold tags are then dissolved and quantified by a simple and sensitive luminol CL reaction. The proposed CL protocol is evaluated for a 30-base model DNA sequence, and the amount as low as 0.01 pmol of DNA is determined, which exhibits a 150 x enhancement in sensitivity over previous gold dissolution-based electrochemical formats and an enhancement of 20 x over the ICPMS detection. Further signal amplification is achieved by the assembly of biotinylated colloidal gold onto the surface of streptavidin-coated polystyrene beads. Such amplified CL transduction allows detection of DNA targets down to the 100 amol level, and offers great promise for ultrasensitive detection of other biorecognition events.  相似文献   

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