荧光免疫夹心法分离及检测EV71病毒

基于免疫磁球分离技术,利用荧光免疫夹心法捕获,建立了一种新的EV71病毒的快速检测方法。用免疫磁球特异性捕获EV71病毒,然后用生物素化的抗体结合EV71病毒,再结合标有量子点的链霉亲和素(SA-QDs)形成复合物,采用荧光光谱法进行定量检测,C4亚型EV71病毒在2.6~7.5lg(TCID50/mL)的范围内线性关系良好(R2=0.9953),检测限为1.7lg(TCID50/mL)。该方法简便快速,灵敏度高。     

基于磁致伸缩材料的猪瘟病毒无线免疫传感器

通过在磁致伸缩材料表面形成金-硫键自组装膜固载猪瘟多抗,构建了一种新型、灵敏、便携、低成本的猪瘟病毒无线免疫传感器。该传感器基于抗原抗体特异性结合使猪瘟病毒附着在磁致伸缩材料表面从而导致磁弹片共振频率降低的原理,通过测定共振频率偏移量实现对猪瘟病毒的无线检测。采用扫描电子显微镜(SEM)对抗体修饰及猪瘟病毒检测过程进行了表征,利用该传感器对不同浓度的猪瘟病毒进行检测,线性范围为0.5~2.5μg/m L,响应灵敏度为65 Hz/(μg·m L~(-1)),检出限为0.5μg/m L。     

聚多巴胺仿生纳米微球用于构建电化学免疫传感器

利用多巴胺仿生聚合方法制备了具有良好生物相容性的聚多巴胺纳米微球,并在其表面原位合成银纳米颗粒.复合物微球具有良好的催化还原H2O2的性能以及良好的结合生物分子的能力.将制备的复合物微球作为标记物,将氨基化石墨烯作为基底材料,构建了检测人免疫球蛋白(Ig G)的夹心型电化学免疫传感器.运用循环伏安法和计时电流法对构建的电化学免疫传感器进行了性能分析,并对实验条件进行了考察优化.在最佳的实验条件下,免疫传感器的线性范围是0.1 pg/m L~15 ng/m L,检出限为0.025 pg/m L.     

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。     

基于Au纳米颗粒/还原氧化石墨烯C-反应蛋白免疫传感器的研制

采用还原氧化石墨烯-金纳米颗粒(RGO-Au NPs)作为免疫传感器的固定基质,将C-反应蛋白(CRP)抗体固定在玻碳电极表面,用蒽醌二羧酸作为标记物,制成夹心型的CRP免疫传感器。在最优实验条件下,通过示差脉冲伏安法对CRP的含量进行检测。该传感器在0.25~100 ng/m L范围内具有良好的线性关系,检出限为0.08 ng/m L,线性系数为0.997。该传感器为C-反应蛋白的检测提供了一种新的手段。     

微流控芯片中磁免疫荧光法检测新城疫病毒

建立了一种基于微流控芯片的磁免疫荧光方法,并用于新城疫病毒(NDV)的高灵敏检测。该方法基于抗原抗体免疫识别作用,采用免疫磁珠实现了对NDV的高效捕获和分离,并利用量子点的荧光作为检出信号,提高方法的灵敏度。在优化条件下,NDV的检测限为1ng/mL,线性范围为1～10ng/mL。利用免疫磁珠捕获分离病毒,可用于家禽肝脏、肺、粪便等实际样品中病毒的检测。该方法可以在1h内完成对NDV的检测,操作简单、特异性强,灵敏度高,重现性好。本文为NDV的检测提供了一种快速、准确的方法。     

基于HRP直接标记的流感病毒H1N1电化学免疫传感器

构建了一种新型、灵敏、便捷式流感病毒免疫传感器,通过在金电极表面键合抗-HA单克隆抗体,选择性捕获目标H1N1流感病毒。该方法基于吸附在病毒表面的辣根过氧化物酶(HRP)在亚甲基蓝(MB)溶液中可有效催化还原H2O2,利用方波伏安法(SWV)考察了还原峰电流的变化,从而实现了对抗原病毒的特异性识别。实验表明所构建的免疫传感器对H2O2-MB体系表现出快速的电流响应以及良好的稳定性,对流感病毒H1N1检测的动态响应范围为0.01~2.0μg/m L,检出限(S/N=3)为5.0 ng/m L。结果证明HRP可作为一种简单快速的探针用于流感病毒的实时监测。     

背景荧光猝灭-免疫层析法检测C-反应蛋白

采用背景荧光猝灭-免疫层析法(b FQICA),基于双抗体夹心法原理,将合适浓度的捕获抗体、示踪抗体分别固定在试纸条和微孔内,层析时与样本形成抗体-抗原-抗体的夹心结构,建立了一种操作简便、灵敏度高、抗干扰性强且能快速定量检测C-反应蛋白(CRP)的方法.结果表明,CRP在0.0~100.0 ng/m L浓度范围内与相对荧光强度值(F1/F2)呈现良好的相关性,最低检出限为0.0939 ng/m L,加标回收率为87.69%~111.0%,检测3批试剂的批间和批内相对标准偏差均小于15%,与降钙素原(PCT,20.0 ng/m L)及人血清淀粉样蛋白A(SAA,10.0μg/m L)均无交叉反应.采用该方法与免疫透射比浊法同时测定41例临床血清样本,检测结果相关性良好(r=0.9585,P0.01),2种方法无显著性差异(P0.05).     

基于免疫磁分离和酶催化放大的化学发光免疫传感器测定人血清中癌胚抗原的含量北大核心CSCD

基于免疫磁分离和酶催化放大策略,构建了一种用于测定人血清中癌胚肮原(CEA)含量的化学发光免疫传感器。利用抗原-抗体的特异性结合,将磁球作为免疫反应的固相载体去捕获癌胚抗原(CEA),并通过在免疫磁球表面负载辣根过氧化酶(HRP)作为检测标记物,形成磁性免疫夹心复合物。通过HRP对过氧化氢的催化作用,氧化鲁米诺产生较强的化学发光信号,根据化学发光强度来定量CEA。结果显示:CEA的质量浓度在0.05~80.00μg·L^(-1)内与其对应的化学发光强度呈线性关系,检出限为4.5 ng·L^(-1);对样品进行回收试验,回收率为97.0%~100%。与其他固相载体和化学发光免疫传感器相比,该方法检测CEA的化学发光强度更强,并且具有更宽的线性范围和更低的检出限。     

免疫磁分离技术结合阻抗测量法快速检测禽流感病毒

采用免疫磁分离和阻抗测量技术联用的方法,实现了对禽流感H5亚型病毒的特异性快速检测。将偶联生物素的H5抗体固定于链霉亲和素修饰的纳米磁珠表面,制备成免疫磁珠,用于样品中禽流感H5N1病毒的分离和浓缩。使用金叉指阵列微电极测量样品阻抗,在特征频率(100 kHz)下,阻抗模值随样品中H5N1病毒浓度增加而增大。研究了抗体浓度、免疫反应时间和磁分离时间对阻抗信号的影响,结果表明,在优化的实验条件(抗体浓度0.25 g/L、免疫反应时间60 min、磁分离时间3 min)下,纯病毒样品和拭子样品中H5N1病毒浓度分别在2!1～24HA unit/50μL和20～24HA unit/50μL范围时,病毒浓度对数值与阻抗信号值呈线性相关关系,检出限分别为0.5 HA unit/50μL和2 HA unit/50μL。     

Determination of pterostilbene in rat plasma by a simple HPLC‐UV method and its application in pre‐clinical pharmacokinetic study

A simple HPLC‐UV method was developed and validated for the quantification of pterostilbene (3,5‐dimethoxy‐4'‐hydroxy‐trans‐stilbene), a pharmacologically active phytoalexin in rat plasma. The assay was carried out by measuring the UV absorbance at 320 nm. Pterostilbene and the internal standard, 3,5,4'‐trimethoxy‐trans‐stilbene eluted at 5.7 and 9.2 min, respectively. The calibration curve (20–2000 ng/mL) was linear (R² > 0.997). The lower limits of detection and of quantification were 6.7 and 20 ng/mL, respectively. The intra‐ and inter‐day precisions in terms of RSD were all lower than 6%. The analytical recovery ranged from 95.5 ± 3.7 to 103.2 ± 0.7% while the absolute recovery ranged from 101.9 ± 1.1 to 104.9 ± 4.4%. This simple HPLC method was subsequently applied in a pharmacokinetic study carried out in Sprague–Dawley rats. The terminal elimination half‐life and clearance of pterostilbene were 96.6 ± 23.7 min and 37.0 ± 2.5 mL/min/kg, respectively, while its absolute oral bioavailability was 12.5 ± 4.7%. Pterostilbene appeared to have better pharmacokinetic characteristics than its natural occurring analog, resveratrol. Copyright © 2009 John Wiley & Sons, Ltd.     

Kinetic spectrophotometric method for the determination of selenium(IV) by its catalytic effect on the reduction of spadns by sulphide in micellar media

A simple kinetic spectrophotometric method was developed for the determination of ultratrace amounts of Se(IV). The method is based on the reduction of spadns by sulphide in micellar media. The reaction was monitored spectrophotometrically by measuring the decrease in the absorbance of spadns at 515 nm with a fixed-time method. The decrease in the absorbance of spadns is proportional to the concentration of Se(IV) in the range 0.5–100 ng/mL with a fixed time of 2.5–7.0 min from the initiation of the reaction. The limit of detection is 0.3 ng/mL Se(IV). The relative standard deviation for the determination of 0.02 and 0.10 μg/mL Se(IV) was 2.10 and 1.95%, respectively. The method was applied to the determination of Se(IV) in water. The text was submitted by the authors in English.     

铑-钨酸盐-罗丹明B缔合显色反应的研究及应用

在聚乙烯醇 (PVA)存在下 ,铑 ( )与钨酸盐及罗丹明 B(RB)形成的离子缔合物在 570 nm处产生最大吸收。建立了测定铑 ( )的光度分析方法 ,其表观摩尔吸光系数 ε达 1 0 7(L·mol-1·cm-1)数量级。方法的检测范围为 0～ 50ng/2 5m L ,检出下限为 0 .2 9ng/2 5m L。大量存在的常见离子对 Rh( )的测定不干扰。用此方法测定了催化剂及工业产品中铑的含量 ,结果与标准方法 (Sn Cl2法 )一致。     

Determination Of Formaldehyde And Acetaldehyde Associated To Atmospheric Aerosols By HPLC

Abstract

A rapid new analytical protocol was developed for the determination of formaldehyde and acetaldehyde associated to atmospheric particulate matter, at ng/m³ levels. The aerosols were collected on glass fiber filters (8″×10″) at face velocities ranging from 15 m/min to 23 m/min. Aliquots of 15.4 cm² were sonicated, for 20 min, with 5.0 mL of 0,01% 2,4-dinitrophenylhydrazine (DNPH), 1 % phosphoric acid. The liquid phase was then filtered and the separation and quantification of the corresponding 2,4-dinitrophenylhidrazone (DNPHo) derivatives carried out by reverse phase HPLC. Acetonitrile:water (57:43, v/v) as mobile phase at 1.0 mL/min and absorbance detection at 350 nm and 365 nm for, respectively, formaldehyde-DNPHo (0.04 AUFS) and acetaldehyde-DNPHo (0.01 AUFS) were used. The precision for four different aliquots, from a 8″×10″ glass fiber filter, were under 0.04% for formaldehyde and 14.16 % for acetaldehyde. In Salvador, Bahia, Brazil, formaldehyde and acetaldehyde were determined, respectively, in the range of 6.8 ng/m³ to 27.3 ng/m³ and 9.1 ng/m³ to 54.6 ng/m³.     

Kinetic-Spectrophotometric Determination of Tellurium (IV) by Its Catalytic Effect on the Reduction of Thionine by Sodium Sulfide in Cationic Micellar Medium

A simple and highly sensitive method is proposed for the determination of Te(IV) by its catalytic effect on the reduction of thionine by sodium sulfide in the presence of cetyl trimethyl ammonium bromide as a micelle media. The reaction was monitored spectrophotometrically by measuring the decrease in absorbance of thionine at 600 nm with a fixed time of 0.5-2.5 min from initiation of the reaction. Tellurium can be measured in the concentration range of 0.6-500.0 ng/mL, with a limit of detection of 0.3 ng/mL Te(IV). The influence of more than thirty potential interfering ions was studied on the determination of tellurium. The relative standard deviation for ten replicate measurements of 0.020 and 50.0 ng/mL Te(IV) was 2.1 and 1.9%, respectively. The method was applied for the determination of tellurium in synthetic samples.     

HPLC of trazodone in serum after microscale protein precipitation

Trazodone, an anti-depressant medication, is found in serum in the 500-1000 ng/mL range in patients taking therapeutic doses. Because of this relatively high concentration, it has been possible to devise an HPLC assay system using the rapid, convenient microscale procedure described previously by Lam et al. (Clin. Chem. 26, 963 1980) to prepare the sample for chromatography. To 0.1 mL serum were added 0.1 mL acetonitrile and 10 microL of 10% zinc sulfate in water. The mixture was centrifuged and 50 microL of the clear supernatant was injected into a reversed-phase column which was eluted with 65% 0.05 M potassium phosphate-35% acetonitrile, with detection by ultraviolet absorbance at 210 nm. The trazodone elutes in 6 min, clearly resolved from endogenous interferences. The recovery of trazodone added to serum was better than 90%. Peak height was proportional to concentrations in the serum sample from 125 ng/mL to 3000 ng/mL.     

HPLC determination of irbesartan in human plasma: its application to pharmacokinetic studies

A simple and rapid HPLC method using fluorescence detection was developed for determination of irbesartan in human plasma. Sample preparation was accomplished through a simple deproteinization procedure with 0.4 mL of acetonitrile containing 800 ng/mL of losartan (internal standard), and to a 0.1 mL plasma sample. Chromatographic separation was performed on a Zorbax Xclipse XDB C₁₈ column (150 × 4.6 mm, i.d., 5 µm) at 40°C. An isocratic mobile phase, acetonitrile:0.1% formic acid (37:63, v/v), was run at a flow‐rate of 1.0 mL/min, and the column eluent was monitored using a fluorescence detector set at excitation and emission wavelengths of 250 and 370 nm, respectively. The retention times of irbesartan and losartan were 4.4 and 5.9 min, respectively. This assay was linear over a concentration range of 10–5000 ng/mL with a lower limit of quantification of 10 ng/mL. The coefficient of variation for this assay precision was less than 8.48%, and the accuracy exceeded 94.4%. The mean relative recoveries of irbesartan and losartan were 98.4 and 99.1%, respectively. This method was successfully applied for pharmacokinetic study after oral administration of irbesartan (300 mg) to 23 Korean healthy male volunteers. Copyright © 2009 John Wiley & Sons, Ltd.     

免疫金铂纳米合金催化共振散射光谱法测定痕量人绒毛膜促性腺激素

以NaBH₄为还原剂, 制备了金与铂物质的量比为49∶1的金铂纳米合金(GP). 用兔抗人绒毛膜促性腺激素抗体(RhCG)修饰AuPt获得了免疫纳米合金探针(GP-RhCG). 在pH 5.8磷酸氢二钠-柠檬酸缓冲溶液及KCl存在的条件下, GP-RhCG探针发生非特异性聚集, 在590 nm处有一个较强的共振散射峰. 当有人绒毛膜促性腺激素(hCG)存在时, 聚集的GP-RhCG探针与hCG发生特异性结合, 生成分散性较好的GP-RhCG-hCG免疫复合物, 导致590 nm处共振散射峰强度降低. 其共振散射峰强度降低值ΔI_{590 nm}与hCG浓度在6.67～86.7 ng/mL范围内呈现良好线性关系. 免疫反应液中形成的GP-RhCG-hCG免疫复合物对葡萄糖-铜(II)体系具有较强的催化作用, 其产物在610 nm处有一较强共振散射峰. 随着hCG浓度增大, 形成的GP-RhCG-hCG复合物越多, 其催化作用增强, 610 nm处的共振散射峰增强. 其共振散射峰增大值ΔI_{610 nm}与hCG浓度在3.33～133 ng/mL范围内呈线性关系.     

合成的近红外花菁染料测定痕量血清蛋白质

染料结合法测定蛋白质在生化及临床分析中已有广泛应用,常用的染料有溴甲酚绿、溴酚蓝、考马斯亮蓝及铬天青S等,这些染料的最大吸收波长均处于可见光区.近年来,近红外染料越来越引起人们的注意.     

Speciation of vanadium (IV) and vanadium (V) with Eriochrome Cyanine R in natural waters by solid phase spectrophotometry

The separation and preconcentration of vanadium (IV) and vanadium (V) using Sephadex DEAE A-25 with Eriochrome Cyanine R has been studied, based on the preconcentration of vanadium (IV) in the first step and V(V) after reduction with ascorbic acid in the second step. Factors affecting the optimum fixation of the complex were investigated. The absorbance of the solid phase is measured directly at 563 nm for V(IV), at 585 nm for V(V) and at 750 nm for both. The proposed method provides a simple and specific procedure for the separation of vanadium in natural waters. The calibration graph is linear up to 150 ng/mL, with RSD of 4.7% for V(IV) and 4.0% for V(V). The detection limits are 1.6 and 1.4 ng/mL for V(IV) and V(V), respectively.