基于固定化酶的化学发光停流法测定D-氨基酸

基于固定化酶的化学发光停流法测定Ｄ－氨基酸封满良,黄玉文,宫志龙,章竹君（陕西师范大学化学系,西安,７１００６２）关键词固定化酶,化学发光,甲壳质,Ｄ－氨基酸自从发现人体有关组织及血桨中Ｄ－氨基酸的水平与某些疾病有关以来,已报道了许多测定Ｄ－氨基酸的...     

稀土Yb(Ⅲ)对Luminol-H2O2-Cr(Ⅲ)体系的化学发光熄灭效应

研究了稀土Ｙｂ（Ⅲ）对Ｌｕｍｉｎｏｌ－Ｈ２Ｏ２－Ｃｒ（Ⅲ）体系化学发光的熄灭效应，在此基础上建立了稀土Ｙｂ的流动注射化学发光分析法。该法灵敏度高，线性范围宽，仪器设备简单，操作方便     

增强化学发光酶免疫分析法测定血清中的铁蛋白

本文采用对碘苯酚增强的Ｌｕｍｉｎｏｌ－Ｈ２Ｏ２－ＨＲＰ化学发光反应体系作为免疫分析的最终检测手段，建立了一种新的铁蛋白的免疫分析方法，与酶联免疫分析法相比，该方法具有灵敏度高，线性范围宽等特点。方法的相对标准偏差３．７％，标准曲线线性范围为０．１２－２０００ｎｇ／ｍｌ，用该方法对血清中的铁蛋白进行了测定，其结果与ＥＬＩＳＡ法所得结果一致。     

高灵敏反相流动注射化学发光法测定抗坏血酸

基于抗坏血酸对Ｆｅ２＋－Ｌｕｍｉｎｏｌ－Ｏ２体系化学发光的抑制效应，利用反相流动注射技术，建立了一个测定抗坏血酸的高灵敏化学发光新方法。该法检出限达２×１０－１０ｇ／ｍＬ；抗坏血酸浓度在 １×１０－９～１× １０－６ｇ／ｍＬ范围内与体系化学发光强度减小量呈良好线性相关；相对标准偏差为１．９％（Ｃ＝５×１０－８ｇ／ｍＬ，ｎ＝１１）。     

一种测定微量碘离子的简便偶合化学发光法

报道了在含Ｉ＾－的Ａｓ箩丹明Ｂ混合溶液中直接注射Ｃｅ＾４－，通过记录化学发光持续时间对微量Ｉ＾－进行定量的偶合化学发光法。检出限为６．０×１０＾－８ｍｏｌ／Ｌ，线性响应的浓度范围为１．０×１０＾－７－１．０×１０＾－５ｍｏｌ／Ｌ。该法简便易行，所受的干扰小，用于加碘食盐，酱油及实际水样中微量Ｉ＾－的测定，结果满意。     

钌(Ⅱ)-联吡啶-抗坏血酸-铈(Ⅳ)体系化学发光测定抗坏血酸

1引言钌（Ⅱ）－联吡啶是一种常用的电致化学发光试剂，我们已成功地用于化学发光测定6-巯基嘌呤。在实验中发现抗坏血酸能增强饰（Ⅳ）氧化钌（Ⅱ）联吡啶的化学发光强度，据此建立了化学发光测定抗坏血酸的新力法。线性范围为3．4×10－8～2．6×10－5mol／L，与文献报道的方法相比扩大了一个数量级，检出限降低约100倍，达1．6×10－8mol／L，且条件温和，重视性好。本法用于药物中抗坏血酸的测定，获得满意结果。2实验部分2．1仪器与试剂LKB-1251型化学发光光度计（LKH公司），配有DispenserSVD自动进样器和DispenserColltl、l卜r…     

邻硝基苯基荧光酮化学发光性能和机理研究

研究了在碱性介质中金属离子催化邻硝基苯基荧光酮（ｏ－ＮＰＦ）－Ｈ２Ｏ２体系的化学发光行为。结果表明，在研究的２０多种金属离子中，只有Ｃｏ２＋离子对ｏ－ＮＰＦ－Ｈ２Ｏ２体系有较强的催化发光作用，Ｃｏ２＋的浓度与发光强度在２．５×１０－９～３．０×１０－６ｇ／ｍＬ范围内呈良好的线性关系。通过对化学发光光谱、荧光光谱的研究以及用Ｈｕｃｋｅｌ分子轨道法（ＨＭＯ）计算了ｏ－ＮＰＦ的π电子分布和键级，提出了ｏ－ＮＰＦ可能的化学发光机理．     

高碘酸钠-过氧化氢-盐酸黄连素化学发光体系的研究

１引言 盐酸黄连素是一种用于治疗肠道感染，毒性低、副作用小的天然药物。对盐酸黄连素的测定，化学发光法未见报道。本文发现，过氧化氢存在下，磷酸介质中盐酸黄连素与高碘酸钠反应产生强的化学发光。据此结合流动注射技术建立了在ＮａＩＯ４－Ｈ３ＰＯ４－Ｈ２Ｏ２－盐酸黄连素体系中测定盐酸黄连素的化学发光新方法。方法的检出限为 ３．８×１０－９ｇ／ｍＬ。线性范围是 １．０× １０－８－ ７．０ ×１０－６ｇ／ｍＬ。相对标准偏差为 ２．３％（ Ｃ８＝ １． ０ ×１０－６ｇ／ｍＬ； ｎ＝１１）。该法与药典方法对照，结果满意。…     

高灵敏反应流动注射化学发光法测定抗坏血酸

基于抗坏血酸对Ｆｅ＾２＋－Ｌｕｍｉｎｏｌ－Ｏ２体系化学光的抑制效应，利用反应相流注射技术，建立了一个测定抗坏血酸的高灵敏化学发光新方法。该法检出限达２＊１０＾－１０ｇ／ｍＬ；抗坏血酸浓度在１＊１０＾－９－１＊１０＾－６ｇ／ｍＬ范围内与体系化学发光强度减小量呈良好线性相关；相对标准偏差为１．９％。     

聚乙二醇辛基苯基醚—铁（Ⅱ）—过氧化氢化学发光体系研究及应用

本报道了聚乙二醇辛基苯基醚（ＯＰ）－Ｆｅ（Ⅱ）－Ｈ２Ｏ２化学发光体系，建立了高等反悔一测定微量铁的化学发光新方法，检出限为７．６×１０＾－８ｍｏｌ／Ｌ，线性响应的浓度范围为１．０×１０＾－７－１．０×１０＾－３ｍｏｌ／Ｌ，该法所受金属离子孤干扰小，用于实际水样及铜精矿样品的全铁的测定，结果令满意。     

化学发光酶联免疫分析检测血清中麻疹病毒抗体

化学发光酶联免疫分析检测血清中麻疹病毒抗体章竹君，邹克渭，程明洁（陕西师范大学分析科学研究所，西安，７１００６２）（陕西省卫生防疫站）关键词麻疹病毒抗体，化学发光，酶联免疫分析，辣根过氧化物酶目前在计划免疫工作中通常采用间接酶联免疫吸附分析法（ＥＬＩ...     

红细胞标记抗体的化学发光免疫分析

用红细胞代替辣根过氧化物酶作为双抗体夹心免疫分析中第二抗体的标记物, 建立了一种红细胞标记抗体的免疫化学发光测定乙型肝炎病毒表面抗原的新方法. 在免疫反应完成后, 结合了抗原-抗体免疫复合物的致敏红细胞在低渗溶液中溶血, 释放出血红蛋白. 基于血红蛋白对鲁米诺-H2O2体系化学发光具有催化作用的原理, 采用化学发光法测定血红蛋白含量. 测得的血红蛋白发光强度与待测抗原浓度呈线性关系. 采用这种方法可检测出0.5 ng/mL的乙型肝炎病毒表面抗原. 将该方法与酶联免疫吸附分析(ELISA)结合起来对乙型肝炎患者血清乙肝病毒表面抗原(HBsAg)进行检测, 两者符合率均为97%, 表明本法具有良好的灵敏度和特异性, 可用于临床标本测试.     

pH敏感相分离高分子的免疫分析应用研究

合成了溶解性受pH值影响的高分子化合物聚甲基丙烯酸甲酯-丙烯酸-顺丁烯二酸酐(MAM)及聚甲基丙烯酸一丙烯酰胺-N-羟基琥珀酰亚胺丙烯酸酯(MAN),分别与抗体进行了共价连接,将其用作免疫分析载体,利用其溶解性可调节特征,建立了新的免疫分析方法.以聚甲基丙烯酸-丙烯酰胺-N-羟基琥珀酰亚胺丙烯酸酯为载体,对血清中乙肝表面抗原进行了检测,阴阳性检出结果与常规ELISA方法相符.     

Gold(III) enhanced chemiluminescence immunoassay for detection of antibody against ApxIV of Actinobacillus pleuropneumoniae

We report, for the first time, a chemiluminescence immunoassay (CLIA) method based on AuCl(4)(-)-enhanced luminol chemiluminescence (CL) reaction for the highly sensitive detection of ApxIV antibody of Actinobacillus pleuropneumoniae (APP). The AuCl(4)(-), which was the dissolution product of the gold nanoparticle-rabbit anti-pig IgG conjugate, served as an analyte in the CL reaction for the indirect measurement of antibody against ApxIV. The optimal condition of gold dissolution was composed of a 5.0 x 10(-2) M HCl, 1.5 x 10(-2) M NaCl, and 2.5 x 10(-4) M Br(2) solution. Under the optimal conditions, a good correlation between the relative CL photon counting and the dilution coefficient of serum was obtained in the dilution range of 1:160-1:40 000. Based on the analysis of clinical samples, the results indicated that CLIA had remarkable advantages in terms of reliability and practical use compared with indirect hemagglutination (IHA) and enzyme-linked immunosorbent assays (ELISA). The proposed method provided a new tool for the indirect determination of antibody against ApxIV in pig serum samples and showed great potential for numerous applications in immunoassays.     

Micro-plate chemiluminescence enzyme immunoassay for aflatoxin B1 in agricultural products

In this work, a micro-plate chemiluminescence enzyme immunoassay by antibody-coated for the determination of aflatoxin B1 (AFB1) in agricultural products has been established. Aflatoxin B1 antibody (AFB1-Ab) was adsorbed physically on polystyrene micro-plate hole as solid phase antibody, which took place immunity-reaction between antigen and antibody with AFB1 standard solution or samples by direct competition. Luminol-hydrogen peroxide chemiluminescence system catalyzed by horseradish peroxidase (HRP) with p-iodophenol enhancement was used as signal detecting system. The effects of several factors, including composition and pH of coating solution, dilution ratio and amount of antibody and enzyme labeled antigen, time of antibody-coating, incubation and chemiluminescence reaction, and other relevant variables upon the immunoasaay were studied and optimized. The linear range of proposed method for AFB1 was 0.05-10.0 ng g⁻¹ with a correlative coefficient of −0.9997. The sensitivity of the proposed method was 0.01 ng g⁻¹. The RSDs of intra- and inter-assay were less than 12.2% and 10.0%, respectively. This method has been successfully applied to the evaluation of AFB1 in agricultural products with recoveries of 79.8%, 101.9% and 115.4% for low, middle and high concentration samples, respectively. It shows a good correlation with the commercial available ELISA kit for AFB1 with correlative coefficient of 0.9098 indicating that the established CLEIA method can be used to determine AFB1 in real samples.     

CE-based simultaneous liquid-phase noncompetitive enzyme immunoassay for three tumor markers in human serum using electrochemical detection

A method for indirectly detecting horseradish peroxidase (HRP) was described by CE with electrochemical detection. Details of selection for optimum conditions were presented. The detection limit of free HRP was 1.09 x 10(-12) M or 0.94 zmol (S/N = 3). A novel CE-based liquid-phase binding noncompetitive enzyme immunoassay (CE-EIA) was developed. In this method, after the noncompetitive immunoreaction in liquid phase, the free enzyme (HRP)-labeled antibody (Ab*) and the bound enzyme-labeled complex (Ag-Ab*) were separated and then the system of HRP catalyzing H(2)O(2)/o-aminophenol (OAP) reaction was adopted. Prostate specific antigen (PSA), carcinoembryonic antigen (CEA), and human chorionic gonadotropin (HCG) in human serum samples were detected without any sample preparation, with the detection limits (S/N = 3) of 0.22, 0.17 and 0.30 ng/mL, respectively. This technique has been successfully applied to detect simultaneously PSA, CEA, and HCG in 12 min, upon adding these three antigens into human serum to simulate patient serum. It proves that the CE-EIA technique proposed could be developed into a sensitive and new method for simultaneous clinical assay of multianalytes.     

A highly sensitive capillary electrophoresis immunoassay strategy based on dual‐labeled gold nanoparticles enhancing chemiluminescence for the detection of prostate‐specific antigen

An enzyme and antibody dual labeled gold nanoparticles enhancing chemiluminescence strategy was developed for highly sensitive CE immunoassay (IA) of prostate‐specific antigen (PSA). In this work, gold nanoparticles were labeled with horseradish peroxidase and antiprostate specific antigen‐antibody, and used as the marker (Ab^*). After PSA (antigen, Ag) was added into the system, a noncompetitive immune reaction was happen between Ab^* and Ag to form an immune complex (Ag–Ab^*). Subsequently, the obtained Ag–Ab^* and unreacted Ab^* were separated by CE, and the chemiluminescence intensity of Ag‐Ab^* was used to estimate PSA concentration. The calibration curve showed a good linearity in the range of 0.25–10 ng/mL. Based on a S/N of 3, the detection limit for PAS was estimated to be 0.092 ng/mL. Proposed CE method was applied for PSA quantification in human serum samples from healthy volunteers and patients with prostate cancer. The obtained results demonstrated that the proposed CE method may serve as an alternative tool for clinical analysis of PSA.     

Competitive Chemiluminescent Immunoassay for the Sensitive Point-of-Care Determination of Metoprolol

A competitive chemiluminescence immunoassay which can be used for point-of-care testing and blood screening of metoprolol is reported. Four haptens for metoprolol were synthesized. An octanedioic acid-modified hapten was conjugated with bovine serum albumin to serve as the immunogen and the haptens were conjugated with ovalbumin for the coating antigen. Polyclonal antibodies for metoprolol were produced and the detection conditions were optimized. A competitive chemiluminescence immunoassay was established based on the produced antibody with potential for bedside therapeutic monitoring of metoprolol. The limit of detection in phosphate-buffered saline was 2?ng?mL^?1. Satisfactory recovery values from 89.3 to 107.6% in plasma were achieved. The results provided by the reported method were consistent with values obtained by high-performance liquid chromatography for pharmacokinetic studies. The immunoassay has potential to be developed as a test kit offering a simple and cost-effective approach for on-site monitoring of metoprolol.     

Flow Injection Chemiluminescence for the Determination of Estriol via a Noncompetitive Enzyme Immunoassay

A novel approach to the detection of estriol using a flow injection system coupled to enhanced chemiluminescent immunoassay was developed based on noncompetitive immunoassay formats. A conjugated estriol-ovalbumin immobilized immunoaffinity column was inserted into the flow system to trap the unbound horseradish peroxidase (HRP)-labeled antibody after an off-line incubation of estriol and HRP-labeled anti-estriol antibody. The trapped enzyme conjugate was detected by the injection of chemiluminescent substrates to produce enhanced chemiluminescence. The linear range for the determination of estriol is 10.0 to 400 ng · mL⁻¹ with a correlation coefficient of 0.996 and a detection limit of 5.0 ng · mL⁻¹. The total time for sampling and chemiluminescent detection of one sample is 400 seconds after 30 min of pre-incubation. The results for pregnancy serum samples obtained by this method are in good agreement with those obtained using ELISA.     

Development of a sensitive micro-magnetic chemiluminescence enzyme immunoassay for the determination of carcinoembryonic antigen

A micro-magnetic chemiluminescence (CL) enzyme immunoassay with high sensitivity, selectivity, and reproducibility was developed for the determination of the tumor marker, carcinoembryonic antigen (CEA) in human serum. A sandwich scheme assay has been utilized with fluorescein isothiocyanate antibody (FITC)-labeled anti-CEA antibody and alkaline phosphate (ALP)-labeled anti-CEA antibody being used in the CL detection. The CL signal produced by the emission of photons from 4-methoxy-4-(3-phosphate-phenyl)-spiro-(1,2-dioxetane-3,2′-adamantane) (AMPPD) was directly proportional to the amount of analyte present in a sample solution. The influences of the reaction time of antigen with antibody, the reaction time of substrate with label, the dilution ratio of ALP-labeled anti-CEA antibody, the concentration of FITC-labeled anti-CEA antibody, and other relevant variables upon the CL signal were examined and optimized. The CL responses depended linearly on the CEA concentration over the range from 2 to 162 ng mL⁻¹ in a logarithmic plot. Assay sensitivity as low as 0.69 ng mL⁻¹ was achieved. A coefficient of variance of less than 13% was obtained for intra- and inter-assay precision. This method has been successfully applied to the analysis of CEA in human serum. According to the procedure based on spiked standards, the recoveries obtained were 80–110%. Comparison experiments were carried out with the commercially available CEA chemiluminescence immunoassay. Satisfactory results were obtained according to a paired t-test method (t value < t _critical at the 95% confidence level).