基于CdtB的副猪嗜血杆菌病间接ELISA检测方法的建立

发布时间：2018-01-04 16:29

本文关键词：基于CdtB的副猪嗜血杆菌病间接ELISA检测方法的建立　出处：《黑龙江八一农垦大学》2016年硕士论文　论文类型：学位论文

更多相关文章： 副猪嗜血杆菌 rCdtB 间接ELISA

【摘要】：为建立一种快速检测副猪嗜血杆菌的血清学方法,本实验采用重组表达的H.parasuis细胞致死膨胀毒素B亚基蛋白(rCdtB)作为包被抗原,并对各反应条件进行优化,建立了检测H.parasuis血清抗体的间接ELISA方法。以该方法检测猪传染性胃肠炎病毒、猪伪狂犬病病毒、圆环病毒2型、猪瘟病毒、猪病毒性腹泻病毒、猪蓝耳病病毒、大肠杆菌、猪链球菌2型、多杀性巴氏杆菌、胸膜肺炎放线杆菌和H.parasuis阳性血清,除了H.parasuis阳性血清呈阳性外,其余均为阴性,表明该方法特异性良好。批内重复和批间重复的变异系数分别为小于7%和8%。此外,将rCdt B-ELISA方法与商品化试剂盒BioChek HPS-OppA间接ELISA抗体检测方法对背景明确的116份H.parasuis阳性血清和112份H.parasuis阴性血清进行检测,商品化试剂盒阳性符合率为25%,阴性血清符合率为100%,总符合率为62%,其中rCdtB-ELISA方法对阳性血清的检出率高于商品化试剂盒。应用rCdt B-ELISA方法检测330份临床猪血清样品,结果显示,副猪嗜血杆菌抗体阳性检出率为11.8%。应用该方法对H.parasuis 7份阳性血清及4份阴性血清进行保质期试验,结果发现6个月后S/P值均大于0.2417,阴性血清的S/P值均小于0.2417,证明rCdtB-ELISA试剂盒至少可以保存6个月。
[Abstract]:In order to establish a rapid serological method for the detection of Haemophilus parapleus, the recombinant H. parasuis cell line lethal expansion toxin B subunit protein (rCdtB) was used as the coating antigen. The indirect ELISA method for detection of H. parasuis serum antibody was established. The method was used to detect porcine infectious gastroenteritis virus and porcine pseudorabies virus. Circovirus type 2, swine fever virus, porcine viral diarrhea virus, porcine blue ear disease virus, Escherichia coli, Streptococcus suis type 2, Pasteurella multocida. The positive sera of Actinobacillus pleuropneumoniae and H. parasuis were negative except H. parasuis positive serum. The results showed that the specificity of the method was good. The coefficients of variation of intra-and inter-assay repeats were less than 7% and 8, respectively. RCdt B-ELISA and Commercial Kit BioChek. HPS-OppA indirect ELISA antibody assay was used to detect 116 H. parasuis positive sera and 112 H. parasuis negative sera. The positive coincidence rate of commercial kit and negative serum were 25 and 100 respectively, and the total coincidence rate was 62%. The detection rate of positive serum by rCdtB-ELISA method was higher than that by commercial kit. RCdtB-ELISA method was used to detect 330 clinical pig serum samples. The positive rate of Haemophilus parasuis antibody was 11.8%. This method was used to test the shelf life of 7 positive sera and 4 negative sera of H. parasuis. The results showed that after 6 months, the values of S / P were more than 0.2417, and those of negative serum were less than 0.2417. It was proved that the rCdtB-ELISA kit could be kept for at least 6 months.
【学位授予单位】：黑龙江八一农垦大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S855.12

【参考文献】