微流控芯片直接化学发光检测方法研究

基于Ru(bipy)₃²⁺的化学发光反应,建立了一种新型微流控芯片化学发光检测方法.以草酸为研究对象,探讨了表面活性剂CTAB的浓度、化学发光反应酸度以及Ru(bipy)₃²⁺浓度等因素对峰形以及检测灵敏度和重现性的影响.草酸的线性范围为1.0×10^-4~5.0×10^-6mol/L,检测限达到2.0×10^-6mol/L.该方法的建立为微流控芯片分离检测某些有机酸、药物、环境物质及核酸等提供了新方法.     

微流控芯片化学发光检测系统研究

在微流控芯片上实现了鲁米诺-过氧化氢-Co2+化学发光反应及分析应用研究。探讨了分离电压对电泳图谱的影响,发现在选定实验条件下,Co2+检出限可达到2.0×10-6mol/L;并且在微流控芯片上实现了Co2+与Cu2+的快速分离及检测。     

微流控芯片化学发光检测系统的设计及其应用

通过标准光刻、化学刻蚀及热键合技术制作微流控电泳芯片,在芯片上集成流通式化学发光检测池,实现样品的芯片电泳分离化学发光检测.采用双(2,4,6-三氣苯基)草酸酯(TCPO)-过氧化氢化学发光体系,通过微泵输送化学发光试剂.单酰化苯并氨基酸和单酰化肌氨酸在该系统中得以成功地分离检测,其检测限分别达到2.8和3.2 μmol/L.     

化学发光检测在微流控芯片中的应用综述

综述了近年来化学发光检测在微流控芯片中的应用.指出微流控芯片(又称为"芯片实验室"或者"微型全分析系统")因具有小型化、集成化和自动化等特点而在近20年来日益受到关注,而化学发光检测具有仪器结构简单、背景噪音低、操作和维护成本低等优点,非常适合用作微流控芯片的检测手段.     

高灵敏的化学发光微流控电泳芯片检测系统的研究

化学发光检测具有灵敏度高、光学构型简单和背景低等特点,非常适合于毛细管电泳和微流控芯片检测.毛细管电泳-化学发光检测方法已成功地用于氨基酸、蛋白质、ATP、过渡金属离子和镧系元素离子等的检测,对金属离子的检测限达到l0¹²mol/L.     

氯丙嗪分子印迹化学发光微流控传感器芯片的研究

以氯丙嗪分子印迹聚合物为识别物质,以鲁米诺-K3Fe(CN)6化学发光体系,建立了一种新型的氯丙嗪化学发光微流控分子印迹传感器芯片的检测方法。利用二氧化碳激光在聚甲基丙烯酸甲酯材质上刻蚀出200μm宽,150μm深的微通道,8 mm长,1 mm宽,0.5 mm深的微检测池。微检测池中填充50μm粒径大小的热聚合得到的氯丙嗪分子印迹聚合物作为识别物质,在线富集氯丙嗪,富集的氯丙嗪可以增强鲁米诺和K3Fe(CN)6的化学发光强度,以化学发光强度定量氯丙嗪量。该传感器的响应值与0.02~0.4μg/mL氯丙嗪呈良好的线性关系,检出限为8 ng/mL(3σ)。该微流控传感器芯片已用于测定人尿液中的氯丙嗪。     

化学发光二维两点检测微流控芯片系统

化学发光二维两点检测微流控芯片系统设计集成了一种可用于分离检测氨基酸、多肽和蛋白质等复杂样品的化学发光二维两点检测微流控芯片系统.该系统采用双检测器同时检测第一维和第二维的分离峰信息,可获得样品的二维分离谱图,满足了对多种复杂结构微流控芯片分离特性进行研究的要求.     

基于微流控芯片电泳化学发光检测人单个红细胞中血红蛋白的含量

运用自行设计组装的微流控芯片电泳化学发光检测装置和单细胞分析专用玻璃微流控芯片,建立了一种测定人单个血红细胞中血红蛋白(Hb)含量的新方法。该方法采用双T型的窄进样通道,宽反应通道及适中分离通道的玻璃微流控芯片,集成单个细胞的进样、固定、溶胞、分离和检测等操作于一块微流控芯片上。以p H 10.5的硼砂缓冲液为电泳介质,选用鲁米诺-过氧化氢化学发光体系,对人单个红细胞中血红蛋白的含量进行测定。血红蛋白的质量在2.0~90 pg范围内,与化学发光强度(峰高)呈良好的线性关系,检出限(S/N=3)为0.8 pg。通过对19个血红细胞进行检测,得到人单个血红细胞中血红蛋白的含量在14~68 pg范围内,该结果与无氰HGB测量法测得的总体细胞血红蛋白的平均值(34.5 pg)基本一致。     

基于分子印迹识别的化学发光微流控传感器芯片测定双嘧达莫

试验中,以双嘧达莫为模板分子,α-甲基丙烯酸为功能单体,乙二醇二甲基丙烯酸酯为交联剂,氯仿为溶剂,合成了双嘧达莫分子印迹聚合物。将此聚合物填充在长8 mm,宽1 mm,深0.5 mm的微流控芯片检测池中作为分子识别物质,设计了一种新型的化学发光微流控传感器芯片测定双嘧达莫。双嘧达莫被此聚合物在线吸附并识别,被吸附的双嘧达莫与鲁米诺和铁氰化钾混合溶液反应并导致其化学发光强度增大。该传感器对双嘧达莫响应范围为1.0～20μg·L~(-1),检出限(3σ)为0.5μg·L~(-1),对10μg·L~(-1)双嘧达莫连续平行测定7次,其相对标准偏差为4.6%。     

基于电渗驱动的微流控芯片顺序注射分析系统

本文介绍了一种在微流控芯片上依靠电渗驱动完成顺序注射分析的新方法。采用由多个双T通道构成的微通道网络,利用夹流进样和电渗流驱动方法,在通道内形成间隔排列的溶液区带,进而在流动过程中实现溶液区带间的相互混合。实验以荧光素溶液与硼砂缓冲液为模型体系,初步考察了利用该系统实现顺序注射进样及混合的效果。该方法装置简单,操作简便,无需任何复杂机械装置,利用微流控芯片上的微通道网络及电渗流控制,实现了亚纳升级的溶液区带的顺序排列和进样。     

流动注射化学发光抑制法测定吡罗昔康

基于吖啶橙在氢氧化钠介质中能被高锰酸钾氧化产生较强的化学发光,吡罗昔康能强烈抑制其化学发光,建立了高锰酸钾-吖啶橙-吡罗昔康化学发光抑制测定吡罗昔康的新方法。吡罗昔康在1.0×10-5~7.0×10-4g/mL范围内与化学发光强度呈良好线性关系,方法的检出限为4.5×10-6g/mL,对1.0×10-5g/mL吡罗昔康连续进行6次测定的相对标准偏差为3.6%。     

溴酸钾-酸性铬蓝K化学发光法检测水中微量NO2-

利用NO2-对溴酸钾-酸性铬蓝K体系的化学发光有显著的增强作用,建立了检测亚硝酸盐的新方法。着重对化学发光条件,包括溴酸钾浓度、酸性铬蓝K浓度、管长、流速及酸度条件进行了实验研究,并利用该方法对地下水及当地雨水进行了检测。实验结果表明,当溴酸钾浓度为0.1 mol/L、酸性铬蓝K浓度为1.0×10-4mol/L、管长为40 cm、流速为2.5 mL/min及硫酸为0.5 mol/L时,本方法检测NO2-的线性范围为1.0×10-8~8.0×10-5mol/L,检出限为1.1×10-10mol/L;对1.0×10-7mol/L和1.0×10-5mol/L的NO2-进行11次平行测定,相对标准偏差分别为1.6%和2.1%;对天然水中NO2-检测,结果满意。对化学发光机理进行了初步探讨。     

Chemiluminescent Determination of MicroRNA-141 Using Target-Dependent Activation of the Peroxidase-Mimicking DNAzyme

A chemiluminescent method was developed for microRNA-141 (miRNA-141) detection based on the target-dependent activation of peroxidase-mimicking DNAzyme. The structure of the probes was optimized which allowed the development of a sensitive method for miRNA-141. Under the optimized conditions, the detection limit and the linear range were 100?pM and 0.1–50?nM, respectively. The sensitivity of the assay was 270,000?nM^?1. The values of coefficient of variation measured within the working range varied less than 2%, which indicates excellent precision for the proposed method.     

Design of a High-Sensitivity Dimeric G-Quadruplex/Hemin DNAzyme Biosensor for Norovirus Detection

G-quadruplexes can bind with hemin to form peroxidase-like DNAzymes that are widely used in the design of biosensors. However, the catalytic activity of G-quadruplex/hemin DNAzyme is relatively low compared with natural peroxidase, which hampers its sensitivity and, thus, its application in the detection of nucleic acids. In this study, we developed a high-sensitivity biosensor targeting norovirus nucleic acids through rationally introducing a dimeric G-quadruplex structure into the DNAzyme. In this strategy, two separate molecular beacons each having a G-quadruplex-forming sequence embedded in the stem structure are brought together through hybridization with a target DNA strand, and thus forms a three-way junction architecture and allows a dimeric G-quadruplex to form, which, upon binding with hemin, has a synergistic enhancement of catalytic activities. This provides a high-sensitivity colorimetric readout by the catalyzing H₂O₂-mediated oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline -6-sulfonic acid) diammonium salt (ABTS). Up to 10 nM of target DNA can be detected through colorimetric observation with the naked eye using our strategy. Hence, our approach provides a non-amplifying, non-labeling, simple-operating, cost-effective colorimetric biosensing method for target nucleic acids, such as norovirus-conserved sequence detection, and highlights the further implication of higher-order multimerized G-quadruplex structures in the design of high-sensitivity biosensors.     

A Thermophilic Tetramolecular G‐Quadruplex/Hemin DNAzyme

The quadruplex‐based DNAzyme system is one of the most useful artificial enzymes or catalysts; their unique properties make them reliable alternatives to proteins for performing catalytic transformation. The first prototype of a thermally stable DNAzyme system is presented. This thermophilic DNAzyme is capable of oxidizing substrates at high temperatures (up to 95 °C) and long reaction times (up to 18 h at 75 °C). The catalytic activity of the DNAzymes were investigated with the standard peroxidase‐mimicking oxidation of 2,2′‐azino‐bis(3‐ethylbenzothiozoline‐6‐sulfonic acid) (ABTS) by H₂O₂. The step‐by‐step design of this unique heat‐activated G‐quadruplex/hemin catalyst, including the modification of adenines at both ends of G‐tracts, the choice of cation, and its concentration for DNAzyme stabilization, is described. This work investigates thoroughly the molecular basis of these catalytic properties and provides an example of an industrially relevant application.     

Ultrasensitive immunoassay of microcystins-LR using G-quadruplex DNAzyme as an electrocatalyst

An electrochemical immunoassay for microcystin-LR (MC-LR) detection was developed using multi-labeled horseradish peroxidase-mimicking DNAzyme on carbon nanotubes (CNTs) as electrocatalyst for signal amplification. CNTs were covalently conjugated to multiple DNAzyme along with MC-LR for a competitive immunoassay. The as-prepared DNAzyme/CNTs/MC-LR biolabel was specifically captured on the electrode surface, and current responses were obtained upon the electro-catalytic reduction of hydrogen peroxide by the captured biolabels. Under optimal conditions, the electro-catalytic current decreased linearly with the increase amount of MC-LR in the range from 0.01 to 7.0 µg L^?1. The linear regression equation was I (µA) = 12.96 ? 1.48 X _[MC–LR] (µg L^?1), with a correlation coefficient of 0.989. The limit of detection of MC-LR was 2.31 ng L^?1. Application of the immunoassay method and LC/MS/MS method for MC-LR determination on spiked reservoir water gave recovery range of 91.7–105.2% and 94.0–105.0%, respectively. The resulting versatile immunoassay exhibited high sensitivity, good precision and satisfactory reproducibility, which could have vast potential in routine water quality monitoring for various environmental toxins.     

A homogeneous hemin/G-quadruplex DNAzyme based turn-on chemiluminescence aptasensor for interferon-gamma detection via in-situ assembly of luminol functionalized gold nanoparticles,deoxyribonucleic acid,interferon-gamma and hemin

A homogeneous hemin/G-quadruplex DNAzyme (HGDNAzyme) based turn-on chemiluminescence aptasensor for interferon-gamma (IFN-γ) detection is developed, via dynamic in-situ assembly of luminol functionalized gold nanoparticles (lum-AuNPs), DNA, IFN-γ and hemin. The G-quadruplex oligomer of the HGDNAzyme was split into two halves, which was connected with the complementary sequence of P1 (IFN-γ-binding aptamer) to form the oligonucleotide P2. P2 hybridized with IFN-γ-binding aptamer and meanwhile assembled onto lum-AuNPs through biotin–streptavidin specific interaction. When IFN-γ was recognized by aptamer, P2 was released into the solution. The two lateral portions of P2 combined with hemin to yield the catalytic hemin/G-quadruplex DNAzyme, which amplified the luminol oxidation for a turn-on chemiluminescence signaling. Based on this strategy, the homogeneous aptasensor enables the facile detection of IFN-γ in a range of 0.5–100 nM. Moreover, the aptasensor showed high sensitivity (0.4 nM) and satisfactory specificity, pointing to great potential applications in clinical analysis.     

反向流动注射化学发光法测定痕量硫离子

在碱性条件下,基于硫离子对鲁米诺和高碘酸钾化学发光反应有很强的增敏作用,建立了反向流动注射化学发光法检测水中痕量硫离子的新方法。在优化实验条件下,硫离子浓度在8.0×10-10~5.0×10-6g/mL范围内与发光强度呈良好的线性关系;检出限(3σ)为2.0×10-10g/mL;相对标准偏差为2.3%(n=11,1.0×10-8g/mLS2-)。该方法用于环境水样中S2-的测定,结果令人满意。     

检测DL-苹果酸的高锰酸钾化学发光体系

利用高锰酸钾化学发光体系测定了DL-苹果酸 ,测得该物质的浓度与其对应的化学发光峰值在一定范围内存在较好的线性关系。测定DL-苹果酸的线性范围在1.0×10-6～1.0×10-2mol·L-1;相对标准偏差为1.4%(n=8) ,检出限为1.0×10-9mol·L-1。测定实际苹果汁与饮料 ,结果良好。实验表明 ,该法线性范围宽、重现性好、操作简便。文中还探讨了该体系的自催化属性。