高尔基蛋白73(GP73)ELISA定量检测方法的建立及临床应用

目的 建立双抗体夹心ELISA定量检测人血清高尔基蛋白73(GP73)的方法,并用于检测血清GP73含量.方法 利用杂交瘤法制备纯化的抗GP73单克隆抗体(mAb)进行辣根过氧化物酶(HRP)标记后,通过棋盘滴定法确定包被抗体和酶标抗体及其最适工作浓度;以重组人的GP73抗原为标准品建立标准曲线;以重复性、灵敏性和回收率实验评价ELISA方法.应用该ELISA法对正常人、肝炎、肝硬化及肝癌患者血清中的GP73水平进行定量检测.结果 当抗GP73 mAb 3D5A10和7H3F5分别为捕获抗体和酶标抗体、工作浓度分别为40 μg/ml和1∶ 4000时,双抗体夹心ELISA法最优.该方法的平均批内和批间变异系数分别为4.6％和7.6％,灵敏度达3.76 ng/ml,平均回收率为(102.2±5)％.用本方法重复测定肝癌(33例)、肝硬化(28例)、乙型肝炎(44例)及正常人(44例)血清样本中GP73浓度(中位数)分别为25.9、27.6、16.7、9.9μg/L;除肝癌与肝硬化组间差异无统计学意义外,其余各组间均有统计学差异(P＜0.05).结论 成功建立了双抗体夹心ELISA定量检测GP73的方法.检测血清GP73可作为诊断肝脏疾病的重要辅助手段之一.     

急性冠脉综合征患者血清高敏C反应蛋白和组织蛋白酶S水平变化与临床意义

目的：探讨血清高敏C反应蛋白（hs—CRP）、组织蛋白酶S（CatS）与急性冠脉综合征的相关关系。方法：选择53例急性冠脉综合征患者（ACS组），对照组36例，检测两组血清CatS、hs—CRP水平并进行比较。同时检测ACS组治疗后血清hs—CRP、Cats水平的变化。结果：ACS组在性别、年龄、吸烟、糖尿病、脑梗死、高血压患病率方面与对照组相比，差异无统计学意义（P〉0．05）；ACS组治疗前血清CatS水平为（0．51±0．03）nmol／L，hs—CRP为（10．21±5．03）mg／L，分别高于对照组的（0．43±0．04）nmol／L和（3．64±1．87）mg／L，差异具有统计学意义（P〈0．05）；ACS组治疗后血清CatS、hs—CRP水平分别为（0．45±0．06）nmol／L和（6．05±3．25）mg／L，较治疗前的（0．51--4-0．03）nmol／L和（10．21±5．03）mg／L明显降低，差异有统计学意义（P〈0．05）。结论：ACS患者血清CatS及hs—CRP的水平的变化提示CatS及hs—CRP可能在ACS发生发展过程中具有重要作用，推测CatS可能作为ACS的一种新的生物标志物。     

血清C反应蛋白对非体外循环下冠状动脉旁路移植术临床疗效的影响

目的探讨术前血清C反应蛋白水平对非体外循环下冠状动脉旁路移植术临床疗效的影响。方法将734例行非体外循环下冠状动脉旁路移植术的冠心病患者根据术前C反应蛋白水平分为CRP≥3．0mg／L组（386例）和CRP〈3．0mg／L组（348例）,记录术后心血管事件发生情况,Logistic回归分析影响预后的相关因素。结果术后30d内CRP≥3．0mg／L组心血管事件发生率[4．9％（19例）]高于CRP〈3．0mg／L组[4．0％（14例）],但差异无统计学意义（P〉0．05）;平均随访（5．2±1．8）年,CRP≥3．0mg／L组心血管事件发生率[22．0％（78例）]明显高于CRP〈3．0mg／L组[12．2％（40例）],差异有统计学意义（P〈0．05）,Logistic回归分析示：CRP≥3．0mg／L不能预测术后30d心血管事件发生情况,但是预测术后远期心血管事件最强因子,COX分析术前CRP≥3．0mg／L是影响患者预后的危险因素,其次左心室射血分数〈45％。CRP≥．0mg／L组5年实际生存率明显低于CRP〈3．0mg／L组[（90．0±1．2）％比（94．0±0．8）％,P〈0．05]。CRP≥3．0mg／L和左心室射血分数〈45％二者合并的患者5年生存率明显降低（88．0±0．4）％。结论血清C反应蛋白水平可以预测非体外循环下冠状动脉旁路移植术的远期临床疗效,提示血管壁的炎症程度与远期疗效相关。     

血清基质金属蛋白酶-9和C反应蛋白与急性冠脉综合征的相关性

目的 研究血清基质金属蛋白酶-9（MMP-9）和C反应蛋白（CRP）与急性冠脉综合征（ACS）的关系。方法选择42例ACS患者、27例SAP患者及25例健康对照,用ELISA法检测血清MMP-9浓度,免疫散射比浊法测定CRP浓度。结果ACS组MMP-9、CRP浓度为（50．64±7．68）ng／ml和（10．24±3．47）mg／L,明显高于SAP组和正常组,差异有统计学意义（P〈0．05）,而SAP组与正常组MMP-9、CRP浓度相比差异无统计学意义（P〉0．05）。结论急性冠脉综合征患者血清基质金属蛋白酶-9（MMP-9）和C反应蛋白（CRP）明显升高。     

糖尿病肾病患者超敏C反应蛋白与β2-微球蛋白的检测分析

目的探讨血清超敏C反应蛋白（Hs-CRP）和β2-微球蛋白（β2-MG）对糖尿病早期肾损伤的诊断价值。方法对62例糖尿病肾病（DN）患者的血清Hs-CRP和β2-MG含量进行测定,并与正常对照组的含量做比较分析。结果DN组Hs—CRP含量为（5．58±2．77）mg／L、β2-MG含量为（9．63±3．98）mg／L,正常对照组Hs-CRP含量为（1．18±0．71）mg／L,β2-MG含量为（4．61±1．75）mg／L。DN组血清Hs。CRP和β2-MG含量均显著高于正常对照组（P〈0．01）。DN组有42例Hs—CRP异常,异常率为67．7％;有44例β2-MG异常,异常率为71．0％。结论Hs—CRP和β2-MG是诊断2型糖尿病早期肾损伤的敏感指标。     

肾移植受者血清CRP浓度监测的临床意义

应用单向放射免疫扩散法（SRID）动态监测102例肾移植受者血清CRP浓度变化。结果显示：血CRP术后升高，第3天达峰值（22．6±10.4mg／L）。1周内迅速下降，持续处于低值0～18．3mg／L（7．5±6.5mg／L）。急性排斥反应时血CRP急剧升高（61．7±14.6mg／L），排斥逆转或移植肾切除后，CRP迅速下降，其变化与血Cr波动呈正相关（γ＝0．4816）。血CRP在细菌感染时亦明显增高（70．5±24．3mg／L），病毒感染时增高不明显（26.8±5.4mg／L）。     

阿托伐他汀对急性冠脉综合征患者外周血单核细胞合成组织因子和TNF-α的影响

目的：研究阿托伐他汀对急性冠脉综合征患者外周血单核细胞表达TNF-α、组织因子水平及组织因子活性的影响。方法：分离急性冠脉综合征患者外周血单核细胞,分别以不同浓度的阿托伐他汀（0,0．1,1,10μmol／L）干预,共同孵育24h后．用夹心酶联免疫吸附测定法检测细胞培养上清液TNF-α及细胞膜组织因子水平,用逆转录聚合酶联链反应测定它们mRNA的表达,同时用底物发光法检测组织因子的活性。结果：不同浓度的阿托伐他汀（0,0．1,1,10μmol／L）干预后急性冠脉综合征患者外周血单核细胞合成TNF-α分别为（306±40）、（264±54）、（229±24）、（189±44）ng／L,合成组织因子分别为（5．8±1．3）、（4．6±0．8）、（3．7±0．9）、（2．7±0．5）ng／L及组织因子的活性分别为（16．2±3．2）、（7．7±2．8）、（4．3±0．8）、（3．8±0．8）pmol／L（均P〈0．05）。结论：阿托伐他汀呈药物浓度依赖性地减少急性冠脉综合征患者外周血单核细胞合成TNF氓组织因子及组织因子活性。阿托伐他汀通过抗炎、抗血栓形成而在急性冠脉综合征防治中起重要作用。     

阿托伐他汀对老年冠状动脉严重病变患者左心室功能的影响

目的探讨不同剂量阿托伐他汀对老年冠脉严重病变且未进行血运重建患者左心室功能的影响。方法连续入选40例冠状动脉造影证实血管严重病变但不适于血运重建术或不愿行血运重建术的老年患者,完全随机分为小剂量组（10mg／d）20例、大剂量组（40mg／d）20例,随访1年,检测基础条件下、治疗6个月、治疗12个月时脑利钠肽前体（Pro—BNP）水平,超声心动图测定左心室舒张末期内径（LVEDD）、左心室收缩末期内径（LVESD）、左心室射血分数（LVEF）等参数变化,并测定血脂水平、炎症因子（hs-CRP）、ALT、AST、CPK等指标。结果治疗6个月时,2组Pro—BNP水平下降[小剂量组（1204．44±1117．72）pg／ml,大剂量组（727．07±720．26）pg／m1],LVEF水平升高[小剂量组（59．634-5．59）％,大剂量组（62．08±5．77）％]（均P〈0．05）;治疗1年时,2组心功能均显著改善,包括血浆Pro—BNP水平下降[小剂量组（778．55±1316．33）pg／ml,大剂量组（344．89±303．52）pg／ml]、LVEF升高[小剂量组（61．56±6．60）％,大剂量组（63．49±8．17）％]、左心室内径缩小（P〈0．01）,但2组问比较差异无统计学意义。3个月后2组患者血浆hs—CRP水平明显下降[小剂量组（4．68±2．19）mg／L,大剂量组（4．63±1．91）mg／L]（P〈0．05）,治疗6个月和1年时总胆固醇水平、低密度脂蛋白水平、hs—CRP水平显著下降6个月[小剂量组分别为（2．08±0．41）mmol／L、（3．40±2．10）mg／L;大剂量组分别为（3．50±0．53）mmol／L、（1．88±0．49）mmo]／L、（2．31±1．55）mg／L。1年时小剂量组分别为（3．50±0．40）mmol／L、（1．97±0．45）mmol／L、（2．34±1．61）mg／L,大剂量组分别为（3．42±0．54）mmol／L、（1．77±0．44）mmol／L、（1．58±1．17）mg／L]（P〈0．01）。治疗1年,所有患者无肝功能异常,肌酸激酶无明显增高。结论在冠脉严重病变患者中无论应用大剂量、小剂量阿托伐他汀均可以显著降低血浆hs—CRP、Pro—BNP水平,改善左心室功能,并且未见明显不良反应。     

hs—CRP在产后静脉窦血栓形成中的含量变化及作用探讨

目的观察超敏C反应蛋白（hs—CRP）在产后静脉窦血栓形成中的含量变化,并探讨其作用。方法24例产后静脉窦血栓形成患者,于人院后第1天及介入溶栓治疗后3、9d空腹抽取静脉血,测定并观察hs—CRP水平变化。结果人院后第1天血清hs—CRP为（37．2±2．4）mg／L,溶栓后3d为（18．5±1．7）mg／L,溶栓后9d为（5．3±0．7）mg／L。3组比较差异均有统计学意义（P〈0．05和P〈0．01）。结论hs—CRP含量变化为静脉窦血栓形成程度提供了定量信息。     

检测大肠杆菌菌体蛋白的夹心ELISA试剂盒的建立及应用研究

目的  建立特异、敏感的检测大肠杆菌菌体蛋白的夹心ELISA试剂盒。方法  采用大肠杆菌菌体蛋白免疫家兔,制备得到高效价抗菌体蛋白抗血清。将经饱和硫酸铵盐析、阴离子交换柱层析和亲和层析纯化的兔抗大肠杆菌菌体蛋白特异性多克隆抗体作为包被抗体和检测抗体,用生物素标记检测抗体,并加入辣根过氧化物酶（HRP）标记的亲和素。结果　建立的大肠杆菌菌体蛋白夹心ELISA检测方法的敏感性为0.32 μg/L,检测范围为1~100 μg/L,与国际同类商品化试剂盒相当。此法具有良好的稳定性,其批内变异系数小于7.7%,批间变异系数小于6.2%。结论  建立了特异、敏感、稳定的检测大肠杆菌菌体蛋白的夹心ELISA试剂盒,为生物制品中残留大肠杆菌菌体蛋白的质量控制提供了一种重要的检测方法。     

原发性高血压左室肥厚患者妊娠相关血浆蛋白-A和C-反应蛋白的变化

目的：探讨原发性高血压（EH）左室肥厚（LVH）患者血浆妊娠相关蛋白-A（pregnancy-associated plasma protein-A,PAPP-A）和C-反应蛋白（Creactive protein,CRP）的变化。方法：EH患者49例,行心脏彩色超声检查,测量室间隔厚度（IVST）、左室后壁厚度（LVPWT）及左室舒张末内径（LVDD）,计算左室质量指数（LVMI）。根据LVH的诊断标准将EH患者分为LVH＋组和LVH一组。运用双抗体夹心酶联免疫吸附实验（EALAS法）测定血浆PAPP-A浓度和运用双抗体放射免疫法检测血浆高敏CRP（hs-CRP）浓度。结果：LVH＋组PAPP—A及hs-CRP均高于LVH-组[分别为8．28（5．90～30．91）mIU／L vs 4．39（3．02～6．98）n1IU／L和8．58（0．90～35．80）mg／L vs 1．90（0．50～2．23）mg／L,P均〈0．05J。PAPP-A与LVMI及hs-CRP与LVMI的相关分析显示：PAPP-A与LVMI呈显著正相关（r=0．649,P〈0．05）,hs-CRP与LVMI呈显著正相关（r=0．441,P〈0．05）。结论：EH伴LVH患者PAPP-A及CRP较LVH-组显著升高,并且PAPP-A、hs-CRP与LVMI呈显著正相关。PAPP—A与CRP可能参与EH患者LVH的形成。PAPP—A与CRP可能是EH患者合并LVH的预测指标。     

副黏病毒Tianjin株重组血凝素-神经氨酸酶单克隆抗体的制备及在临床检测中的初步应用

目的:建立双抗体夹心ELISA法,调查副黏病毒Tianjin株引起婴幼儿下呼吸道感染情况。方法:用纯化的重组HN片段免疫BALB/c小鼠,按常规方法制备单克隆抗体。以兔抗Tianjin株多克隆抗体为捕获抗体,单抗G7G7E7为检测抗体,建立双抗体夹心ELISA法,检测104例下呼吸道感染患儿支气管肺泡灌洗液(BALF)标本。结果:获得3株能稳定分泌单克隆抗体的杂交瘤细胞株G7H4D3、G7D9G3和G7G7E7。3株单克隆抗体与副黏病毒Tianjin株有较高结合活性,与甲、乙型流感病毒,新城疫病毒(NDV),人副流感病毒(hPIV)1型、3型、肺炎支原体均无交叉反应性。ELISA相加试验和阻断试验表明,3株单抗识别相同或相近表位区域,识别的表位在抗病毒免疫应答中处于免疫优势地位。双抗体夹心ELISA法检测阳性率为1.92%(2/104)。结论:与RT-PCR和间接ELISA法比较,双抗体夹心ELISA法,具有敏感性高、特异性强的优点,适于临床检测使用。副黏病毒Tianjin株是婴幼儿下呼吸道感染的重要致病因子之一。     

Demonstration of a direct capture immunoaffinity separation for C-reactive protein using a capillary-based microfluidic device

C-reactive protein (CRP), a biomarker of inflammation and cardiovascular disease (CVD) risk assessment, was selected as a model antigen to demonstrate a direct labeling/direct capture immunoaffinity separation. The miniaturized device for immunoaffinity chromatography was constructed from two syringe pumps, a gradient mixing microchip, micro-injector with 250nL capillary injection loop, a capillary column, and a diode laser-induced fluorescence detector fitted with a fused-silica capillary flow cell. Monoclonal anti-CRP was biotinylated and attached to 5.0mum streptavidin-coated silica beads to make the solid support for separation columns. CRP in simulated serum matrix was fluorescently labeled in a one-step reaction and directly injected onto the immunoaffinity capillary. The purified antigen was then eluted in an acid gradient and measured. The antibody binding of CRP was evaluated in two physiological buffers, phosphate buffered saline (PBS) and Dulbecco's PBS (DPBS). A quadratic calibration model produced % relative errors of -15.9 to 12.6 for CRP concentration levels ranging from 0.47 to 95.0mug/mL. The accuracy (% difference from nominal) and precision (% relative standard deviation) of replicate injections were within 17.0%. The limit of detection was 57.2ng/mL and chromatographic run times were less than 10min. The instrument design is simple, and potentially portable, while the assay procedure may be modified for other clinically relevant markers by changing the capture antibody.     

Detection of high- and low-affinity antibodies against a human monoclonal antibody using various technology platforms

The effect of monoclonal antibody (mAb) affinity on the detection limit of enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance (SPR), and electrochemiluminescence (ECL) methods was evaluated using a panel of murine mAbs with affinities ranging from 0.057 to 340 nM. M1 and M7 are anti-idiotypic mAbs against a human mAb, ABX10, with dissociation equilibrium constant (KD) values of 0.057 and 7.2 nM, respectively. HP6030 and HP6002 are anti human IgG mAbs with KD values of 30 and 340 nM, respectively. The limit of detection (LOD) for these mAbs was determined using ELISA, SPR, and ECL technologies and was generally correlated with the rank order of their affinities. The LODs for M1, M7, HP6030, and HP6002 by ELISA were 17 +/- 13, 26,000 +/- 9,020, 344,000 +/- 271,000, and 792,000 +/- 1,050,000 ng/ml, respectively. According to an industry-suggested detection limit of 500 ng/ml, the ELISA was not sensitive enough for detecting M7, HP6030, and HP6002, demonstrating its limitation for detection of low- affinity mAbs. The SPR method lowered the LOD for M7 to 3,900 ng/ml, which was above the industry requirement. The ECL method lowered the LOD for all antibodies tested. Importantly, the ECL method lowered the LOD for M7 to 570 +/- 370 ng/ml, which is close to the industry requirement. Since the ECL method had demonstrated a high serum tolerance, its detection capability may be improved by using a higher percentage of serum in the assay matrix. Although a hook effect was observed with ECL methods, the methods could still detect anti-drug antibody (ADA) concentrations greater than 1 mg/ml, which minimizes concerns that high-titer ADA responses could be missed. The results demonstrated the superiority of an ECL method in detecting high- and low- affinity antibodies when compared to the ELISA and SPR methods.     

肝癌组织γ-谷氨酰转移酶单克隆抗体制备及ELISA方法的建立

目的应用纯化的人肝癌组织中蔓陀罗凝集素(DSA)强结合的γ-谷氨酰转移酶(GGT)制备单克隆抗体(McAb),并建立ELISA检测方法。方法应用亲和色谱法纯化肝癌组织中DSA强结合的GGT;采用单克隆抗体技术获得其McAb;protein A-Sepharose亲和色谱法纯化McAb,过碘酸钠法标记HRP法后建立ELISA方法。结果获得5株能特异性识别DSA强结合GGT的McAb细胞株,其中3株细胞所分泌McAb具有较高的特异性,应用所获得McAb进行HRP标记后,建立的ELISA方法,其检测灵敏度为10μg/L,批内及批间平均变异系数为8.9%和11.5%。结论成功制备了DSA强结合的GGT McAb杂交瘤细胞系,并建立了ELISA免疫学检测方法,为临床应用打下基础。     

Reduction of matrix interferences by the combination of chaotropic salt and DMSO in a broadly applicable target-based ELISA for pharmacokinetic studies of therapeutic monoclonal antibodies

Use of a synergistic effect of DMSO together with a chaotropic salt (NaSCN or MgCl₂) allowed to drastically reduce matrix interferences in an ELISA for therapeutic monoclonal antibodies. Optimum combinations were found to be 0.4 M NaSCN together with 10.0% DMSO, and 1.0 M MgCl₂ with 15.0% DMSO. At this optimum combination, quality controls spiked with mAb at 50.0 ng/ml in eighteen individual human sera and plasmas were quantified with an overall accuracy of 102.0%. All of these QCs fulfilled the acceptance criteria of 80.0–120.0% accuracy and precision below 20.0%. The assay was also successfully applied to the quantification of two other mAbs in human serum. Furthermore, the use of the assay was extended to pre-clinical species (cynomolgus monkey and rat serum). Here, the performed validation experiments confirmed the utility of the assay and demonstrated that the assay allowed quantification of mAb from 50.0 ng/ml to 100.0 μg/ml in cynomolgus monkey serum. The method has then been applied to a pharmacokinetic study in cynomolgus monkeys. In summary, this work demonstrates the efficacy of the combination of a chaotropic salt with DMSO to minimize matrix interferences in an ELISA. The robustness thus obtained allowed the successful establishment of a cost effective, target-based ELISA format for use in pharmacokinetic studies, that is easily applicable for the quantification of mAbs in various matrices such as human, cynomolgus monkey or rat serum and plasma.     

Development and evaluation of a direct sandwich enzyme-linked immunosorbent assay for the quantification of guinea-pig immunoglobulin e

The purpose of this study was to develop and evaluate a direct sandwich enzyme-linked immunosorbent assay (ELISA) for the immunoglobulin E (IgE) in serum and plasma from guinea pig using mouse monoclonal antibodies specific for guinea-pig IgE. Mouse monoclonal antibodies were raised against purified IgE protein. The ELISA was performed using a combination of two anti-IgE monoclonal antibodies. One antibody was labeled with horseradish peroxidase (HRP), and the other was coated on polystyrene wells. Purified guinea-pig IgE was used as the standard material. The validity of the ELISA was confirmed by precision, dilution, recovery, and interference tests. The range of detection was 3.1-800 ng of IgE mass per mL of serum and plasma. The intra- and inter-assay coefficients of variation were 4.6% and 5.7%, respectively, or less. The recovery test showed variation only between 92.1% and 111.8%, and the anticoagulants showed noninterference with the IgE assay. The mean serum IgE mass concentration in OVA-sensitized guinea pigs was 29438 ng/mL, and it was 48.6 ng/mL in normal guinea pigs. The present ELISA is useful and practical for specific measurement of the guinea-pig IgE, and it is surmised that it would be suitable for use in allergological and pharmacological research.     

A monoclonal antibody ameliorates local inflammation and osteoporosis by targeting TNF-α and RANKL

This study aimed to generate a monoclonal antibody (mAb) targeting both tumor necrosis factor-α (TNF-α) and receptor activator of NF-κB ligand (RANKL) and to evaluate the therapeutic effects of this antibody on acute inflammation and osteoporosis. We used hybridoma techniques to generate potential mAbs and enzyme-linked immunosorbent assay (ELISA) to determine their specificity. Crystal violet staining was performed to measure the effective dose of the candidate mAbs. The neutralizing effect of the mAbs was evaluated by TNF-α-mediated cytotoxicity and RANKL-induced osteoclastogenesis assays. We further assessed the therapeutic effect of the mAbs in BALB/c mice with carrageenan-induced acute inflammation and ovariectomy-induced osteoporosis. We successfully generated an IgG1 isotype mAb that recognizes human TNF-α and RANKL, which we named 8G12. The 50% effective dose of 8G12 was approximately 1 μg/mL. L929 cells treated with 8G12 exhibited decreased levels of apoptosis (20.04% compared to 63.28% in the positive controls). In addition, treatment with 8G12 inhibited osteoclastogenesis in a dose-dependent manner in vitro. Carrageenan-induced paw edema was significantly reduced in the 8G12-treated mice compared to the positive controls. Treatment with 8G12 also reduced the number of infiltrating leukocytes by more than 50%. The 8G12 treatment not only prevented bone loss but also increased the number, thickness and volume of trabeculae and reduced trabecular separation in ovariectomized mice. Our data suggest that the 8G12 effectively neutralizes the bioactivity of TNF-α and RANKL, ameliorating osteoporosis and inflammation. We therefore propose that 8G12 could be a candidate for generating therapeutic antibodies for treating inflammatory bone diseases.     

双抗体夹心酶联免疫法检测不同样品中的蓖麻毒素

目的应用酶联免疫吸附分析方法（ELISA）检测待测样品中的蓖麻毒素。方法用蛋白G亲和层析柱纯化蓖麻毒素单克隆抗体（4C13,3D74,5E4和5H6）,以3D74及辣根过氧化物酶（HRP）标记的4C13建立的双抗体夹心ELISA对含有蓖麻毒素的多种样品进行检测。结果抗蓖麻毒素的单克隆抗体经亲和层析纯化后具有较高的蛋白纯度,应用HRP标记的4C13与3D74建立双抗体夹心ELISA,对于溶解于磷酸缓冲液中的蓖麻毒素标准品的检测灵敏度可达2．5μg·L^-1;对于土壤、面粉、牛奶、咸菜汁、雪碧、可乐和腐乳汁．中的蓖麻毒素样品检测的灵敏度为2．5-5．0μg·L^-1;与磷酸缓冲液样品相比较,含有相同浓度蓖麻毒素的小鼠和人血清样品ELISA的阳性结果明显减弱．结论双抗体奕心酶联免疫法能够有效用于含有蓖麻毒素样品的检测分析。     

Novel CCR5 monoclonal antibodies with potent and broad-spectrum anti-HIV activities

To identify monoclonal antibodies (mAbs) with high potency and novel recognition sites, more than 25,000 of mouse hybridomas were screened and 4 novel anti-human CCR5 mAbs ROAb12, ROAb13, ROAb14, and ROAb18 showing potent activity in cell-cell fusion (CCF) assay were identified. These mAbs demonstrated potent antiviral activities in both single-cycle HIV infection (IC(50) range: 0.16-4.3 microg/ml) and PBMC viral replication (IC(50) range: 0.02-0.04 microg/ml) assays. These potent antiviral effects were donor-independent. All 4 mAbs were also highly potent in the PhenoSense assay against 29 HIV isolates covering clade A through G. In all antiviral assays, these mAbs showed potency superior to the previously reported mAb 2D7 in side-by-side comparison studies. All 4 mAbs were also fully active against viruses resistant to HIV fusion inhibitor enfuvirtide and CCR5 antagonist maraviroc. Although ROAb12, ROAb14, and ROAb18 inhibited RANTES, MIP1alpha and MIP1beta binding and cell activation, the other novel mAb ROAb13 was inactive in inhibiting cell activation by these three ligands. Furthermore, highly synergistic antiviral effects were found between mAb ROAb13 and 2D7 or ROAb12. In addition, none of these mAbs showed agonist activity or caused internalization of the CCR5 receptor.