基于HRP-H_2O_2-OPDA发光体系的流感病毒H1N1酶联免疫传感器

制备了一种基于HRP-H_2O_2-OPDA发光体系的流感病毒H1N1酶联免疫传感器。利用流感病毒H1N1表面的HA(血球凝集素)蛋白与糖蛋白能有效结合的特性,选择辣根过氧化物酶(HRP)作为生物标记物直接标记在流感病毒的表面,HRP能有效催化H_2O_2-邻苯二胺(OPDA)体系产生灵敏的显色反应和荧光效应。实验对HRP孵化时间、H_2O_2-OPDA催化反应时间等参数进行了优化。结果表明,该酶联免疫传感器对流感病毒H1N1检测的动态响应范围为0.0001~0.5μg/m L,检出限为50 pg/m L(3σ)。其他亚型病毒和干扰蛋白对该传感器的干扰较小,表现出良好的选择性。所构建的方法可为其它流感病毒及其它抗原的检测提供参考。     

四羧基酞菁钴(Ⅲ)/Fe3O4/金胶共固定修饰免疫传感器检测尿样中核基质蛋白22

同时固定四羧基酞菁钴Ⅲ(CoPc)和HRP标记核基质蛋白22抗体(HRP-Ab-NMP22)于自组装在金电极表面的Fe3O4/Au胶上,构建了一种快速测定膀胱肿瘤患者尿液中NMP22抗原(NMP22)含量的安培免疫传感器。CoPc可被用作电极表面HRP的电子传递媒介体。当该传感器在含NMP22尿样的溶液中温育后,NMP22与HRP-Ab-NMP22免疫结合导致HRP的活性中心与CoPc之间的电子传递部分阻碍,使HRP对H2O2电催化还原电流Io降低。ΔIo与NMP22浓度在1.2~200μg/L呈线性关系;检出限为0.5μg/L。该传感器对NMP22响应灵敏,较ELISA法提高了检测速度,有望用于膀胱肿瘤的体外诊断。     

白藜芦醇作辣根过氧化物酶底物的布氏杆菌抗体酶联免疫传感器研究

一种纯天然产物白藜芦醇用作辣根过氧化物酶(HRP)底物。对其化学性质的研究证实,白藜芦醇在空气中较稳定,对HRP、H2O2的电化学响应性能优于传统HRP底物,对人体无毒害。白藜芦醇在HRP催化下可被H2O2氧化成醌,产物醌在电极上于-376 mV处可被还原,其电流的大小与HRP的浓试在一定浓度范围内呈线性相关。将兔布氏杆菌抗原包埋在石墨-石蜡基质中制备了测定兔布氏杆菌抗体的电化学酶联免疫传感器,该传感器测定兔布氏杆菌抗体的线性范围为3×10-4~1.65×10-2g/L;检出限为1×10-4g/L;RSD为4.6%。本方法制备免疫传感器的电化学性能稳定,抗原活性保持良好。     

基于新亚甲蓝为介体的免疫传感器的研究

利用共价键合法,将新亚甲蓝(NMB)与辣根过氧化酶(HRP)标记的羊抗小鼠IgG抗体(Ab)修饰于玻碳电极表面,制成一种新型的电流型免疫传感器。研究了该传感器对H2O2的电化学响应及对小鼠IgG抗原(m IgG)的免疫检测。结果表明NMB作为介体能有效地传递电子,测得电子转移系数为0.77,表观反应速率常数为1.18 s-1。利用传感器对m IgG的检测,线性范围为0.5~3μg/L;检出限为0.028μg/L;相关系数r为0.996。     

巯基丁二胺镍(Ⅱ)自组装单分子层H2O2传感器研究

采用巯基丁二胺镍(Ⅱ)(NiL-SH)自组装单分子层修饰金电极,制得测定H2O2的电流型生物传感器.NiL-SH在电极表面有一对可逆的氧化还原峰.传感器对H2O2的还原显示出快速电催化响应(＜10 s),采用电子扫描电镜(SEM)和光电子能谱(XPS)对电极表面进行了表征,且计算了其自身电化学反应性质.研究了各种因素如pH、工作电位等对传感器响应电流的影响.循环伏安法测定H2O2的线性范围为5.0×10-6～3.0×10-3 mol/L(r=0.999),检出限为2.0×10-6 mol/L.该电极显示出模拟酶特性,测定米氏常数KMapp＝2.78 mmol/L.对传感器的稳定性,灵敏度和选择性进行研究,并应用于实际样品测定.     

以新亚甲蓝为介体的过氧化氢传感器的电化学行为研究

利用共价键合法,将新亚甲蓝(NMB)与辣根过氧化酶(HRP)修饰于玻碳电极表面,制成一种新型的电流型H2O2传感器。探讨了该传感器在0·1mol/L磷酸缓冲溶液(pH=7·0)中的电化学性质。结果表明,NMB作为介体能够有效地在辣根过氧化酶和电极之间传递电子。测得电子转移系数为0·861,表观反应速率常数为1·27s-1。研究了传感器对H2O2的响应及动力学性质,米氏常数为8·27μmol/L,线性响应范围为2·5~100μmol/L。同时研究了pH、缓冲容量及温度等因素对H2O2传感器的影响。     

层层自组装法制备普鲁士蓝修饰电极及对过氧化氢的测定

Fe3+可与电沉积在玻碳电极表面的对氨基苯磺酸发生静电吸附作用并与[Fe(CN)6]4-形成普鲁士蓝(PB),进一步交替重复吸附Fe3+和[Fe(CN)6]4-反应,形成PB晶体.该晶体对还原过氧化氢(H2O2)具有很高的电化学活性.通过循环伏安法、交流阻抗法和计时电流法对传感器进行了电化学表征.研究了该传感器对H2O2的电催化作用,探讨了工作电位,PH值及干扰物质对响应电流的影响.结果表明,在磷酸盐缓冲溶液中(pH =5.4,0.1 mol/L),响应电流与H2O2的浓度在0.97 ～ 32.33 mmol/L范围内具有良好的线性关系,相关系数为0.9993,传感器的响应时间小于5 s,检测限为0.48 mmol/L( S/N为3).     

GCE/DNA/PFS/RTIL/HRP修饰电极的制备及其电催化行为

将辣根过氧化物酶（HRP）固定在室温离子液体（RTIL）/聚二茂铁硅烷（PFS）/DNA复合材料修饰的玻碳电极（GCE）表面,构建了GCE/DNA/PFS/RTIL/HRP修饰电极,详细地研究了该修饰电极的电催化行为,优化了电解质溶液的pH值和RTIL的体积对催化过氧化氢（H2O2）的影响。 电化学实验结果表明,DNA、PFS和RTIL复合膜既为HRP提供了一个生物兼容的微环境;又有效地促进了电子在HRP和电极表面之间的传递。 在最优实验条件下,该修饰电极对H2O2具有快速的催化响应,在2 s内即可达到稳态电流的95%,其响应在3.25 μmol/L~1.47 mmol/L（r=0.999,n=10）和1.86~5.35 mmol/L（r=0.996,n=12）范围内呈良好的线性关系,检出限为0.86 μmol/L。 该传感器灵敏度高、重现性和稳定性好。 此外,该修饰电极还能催化O2还原。     

磁性纳米金共价固定癌胚抗原单克隆抗体的电流型免疫传感器

合成了Fe3O4/Au磁性复合纳米粒子, 在粒子表面通过自组装硫脲分子使表面氨基化, 再用戊二醛共价交联固定癌胚抗原抗体(anti-CEA). 在外加磁场的作用下, 将anti-CEA复合磁性粒子吸附在固体石蜡碳糊电极表面, 制成了新型电流型免疫传感器. 免疫电极在含有癌胚抗原CEA和辣根过氧化物酶标记的癌胚抗原(HRP-CEA)的混合溶液中温育, CEA和HRP-CEA与固定在电极表面的anti-CEA发生竞争反应, 导致HRP对H2O2的催化降解作用的改变, 从而可间接测定CEA. 由于标记的HRP可催化降解H2O2, 导致媒介体间苯二酚浓度改变, 使测定的灵敏度大大提高. 响应电流与CEA质量浓度的对数在2~160 ng/mL的范围内呈线性关系, 检出限为0.57 ng/mL(3σ法). 该免疫传感器具有制作简单、价廉及表面易于更新等特点.     

基于包被包膜糖蛋白抗体纳米金磁性微粒的无试剂安培免疫传感器

在玻碳电极(GCE)表面固定对H2O2有催化还原活性的富马酸二甲酯联吡啶铜(GCE|CuL);再在GCE|CuL表面修饰一层金磁微粒-壳聚糖复合膜(nano Au/Fe3O4/Chit), 进而固定艾滋病毒(HIV)诊断标志物--包膜糖蛋白(gp160)抗体(anti gp160), 由此构建了一类快速检测 gp160的无试剂安培免疫传感器.当该传感器在含gp160溶液中37 ℃下温育30 min后, 传感器表面生成的免疫复合物随gp160浓度的增大而增加, 导致CuL 对H2O2 电催化还原效果降低, 催化电流呈现下降趋势.在PBS溶液(pH 7.0)和-300 mV下, 催化电流的降低值ΔIo与gp160浓度在1～400 μg/L 呈线性关系; 检出限为0.5 μg/L(3σ).研制的免疫传感器检测gp160时, 一步免疫反应即可得结果, 较相同条件下包被gp160抗体的纳米金单分子层修饰电极灵敏度更高, 检测范围更宽, 有望用于艾滋病人血清标志物gp160快速筛测.     

A novel oxidative reaction of 2-nitro-1-phenylpropane with sodium nitrite. A new approach to prepare 1-oximino-1-phenylacetones

A useful method for preparation of 1-oximino-1-phenylacetones via a novel oxidative reaction of 2-nitro-1-phenylpropanes with sodium nitrite was reported.     

A new synthetic route to substituted 1-fluoro-1-cyclopropanecarboxylates

Substituted 6-fluoro-2,4-dioxo-3-azabicyclo[3.1.0]hexane-6-carboxylates were obtained as a mixture of the exo- and endo-isomers by reaction of substituted 6,8-dioxo-2,3,7-triazabicyclo[3.3.0]oct-3-en-4-carboxylates with N-fluoropyridinium tetrafluoroborate.     

A one-step immunochromatographic assay for detecting ginsenosides Rb1 and Rg1

An immunochromatographic strip test has been developed for detecting ginsenosides Rb1 (G-Rb1) and Rg1 (G-Rg1). This qualitative assay system is useful as a rapid screening method for detecting G-Rb1 and G-Rg1 in plants and plant preparations. Our assay is a competitive immunoassay that uses anti-G-Rb1 and anti-G-Rg1 monoclonal antibodies (MAbs) and a detection reagent that contains colloidal gold particles coated with anti-G-Rb1 and anti-G-Rg1 MAbs. Detection limits are 2 g mL^–1 for both G-Rb1 and G-Rg1.     

A concise synthesis of 2-benzoyl-1-indanones and 1-indanones from 2-aryl-1-tetralones

Abstract

Methyl-2-(3-oxo-3-aryl) benzoates derived from acid catalyzed air oxidative fragmentation of 2-aryl-1-tetralones were efficiently undergone intramolecular-Claisen condensation in the presence of potassium tertiary butoxide. The resulting 2-benzoyl-1-indanones formed in two-step ring contractions were further subjected to indium triflate mediated retro-Claisen condensation to get 1-indanones.     

CCl2(A1B1和a3B1)被酮类分子猝灭速率常数的测定

对CCl4/Ar混合气体脉冲直流高压放电产生CCl2自由基,经过约110μs后,再用541.52mm激光将电子基态CC2激励到激发态A1B1(0,4,0)振动态K=0能级上,通过检测激发态CCl2(A)时间分辨荧光信号,测得室温下CC2(A1B1和a3B1)被酮类分子猝灭的实验结果,用所提出的三能级模型分析处理实验数据,获得态分辨速率常数KA和Ka值.     

Purification of Complex of SAV1 with PP1A

SAV1 is a core component involved in the Hippo pathway which can control the organ size via regulating cell proliferation and apoptosis simultaneously. We explored the regulatory mechanism of SAV1. We established the HEK293T cell pool, the cells in which can stably express SAV1 by retroviruses infection and found that SAV1 stable cells reduced the movement of themselves and resulted in multicellular aggregation. We purified SAV1 interacting protein complex using streptavidin resin and subsequently analyzed the digested peptides by high performance liquid chromatography(HPLC)-MS/MS. Results show that about 150 proteins were identified in the complex of SAV1 with protein. TUBA1A, OTUD4, and ATD were identified as proteins interacting with SAV1. Importantly, PP1A, serine/threonine protein phosphatase PP1-alpha 1 catalytic subunit, was also in the top 10 list. The interaction between PP1A and SAV1 was detected by both co-immunoprecipitation(CO-IP) and immunostaining. Our results indicate that PP1A might be the phosphatase of SAV1 and may take part in the regulation of the Hippo pathway.     

单管高灵敏度等温扩增技术快速检测甲型H1N1流感病毒

建立了单管逆转录环介导等温扩增法(RT-LAMP)快速检测甲型H1N1流感病毒的方法.针对甲型H1N1流感病毒的M基因和HA基因的保守区,设计了两组特异性引物,分别用于筛选甲型流感病毒及鉴定甲型H1N1流感病毒.对反应体系中的关键因素进行优化,反应结果可直接通过浊度或者SYBR Green Ⅰ荧光进行判定.本方法最低可...     

CCl_2(A~1B_1,a~3B_1)自由基被烷烃类分子猝灭动力学

用放电- LIF实验装置 ,对 CCl4/Ar混合气体放电产生 CCl_2自由基 ,再用 541.52 nm激光将电子基态 CCl_2激励到激发态 A~B_1(0,4,0)振动能级上 ,通过检测激发态 CCl_2时间分辨荧光信号 ,测得室温下 CCl_2(A~B_1)被烷烃类分子猝灭的实验结果 ,用我们提出的三能级模型分析处理实验数据 ,获得 CCl_2(A~1B_1)态和 CCl_2(a~3B_1)态的碰撞猝灭速率常数 k_A和 k_a值 .     

Concomitant detection of CYP1A1 enzymatic activity and CYP1A1 protein in individual cells of a human urothelial cell line using a bilayer microfluidic device

To understand molecular networking at the cellular level, analyses of processes and effects at the single-cell level are most appropriate. Usual biochemical or molecular biological analyses are based on integrated signals of numerous cells which differ, however, in their expression and activity profiles. Here we show that it is possible to determine different types of properties of individual cells by means of a specifically designed microfluidic device. As part of investigations to characterize the human urothelial cell line 5637 as a potential model system for studies of toxic and carcinogenic effects on urothelial cells, we use this cell line to assign cytochrome P450 activity, and expression of the enzymes involved, to individual cells. It is shown that the cell population is very heterogeneous with respect to the extent and kinetics of CYP1A1-dependent ethoxyresorufin O-deethylase (EROD). This is also true for the cells’ CYP1A1 protein content. With some exceptions, the EROD activity largely coincides with the presence of CYP1A1 protein in the cells. The results obtained with the microfluidic device are promising and open up new perspectives with regard to multi-property determinations in individual cells and to studies focusing on the biochemical and molecular heterogeneity of cells. Figure Formation of fluorescent resorufin from ethoxyresorufin by cytochrome P450 activity in urothelial cells attached within the chamber of a microfluidic device