猪圆环病毒1型抗体IPMA检测方法的建立及应用

Development and application of an immunoperoxidase monolayer assay for detection of antibodies against porcine circovirus type 1

建立了一种用于检测猪圆环病毒1型(PCV1)血清抗体的免疫过氧化物酶单层细胞试验(IPMA)方法。对该方法的反应条件、特异性、敏感性及重复性等进行了系统试验,并与用昆虫杆状病毒表达的PCV1-Cap重组蛋白抗原建立的间接ELISA的检测结果进行了比较。结果显示,用IPMA法对已知PCV1和猪圆环病毒2型(PCV2)参考阳性血清进行检测,血清稀释度≥1∶100即可消除2种病毒间的低水平交叉反应,与其他几种猪病毒参考阳性血清无交叉反应;与用昆虫杆状病毒表达的PCV1-Cap重组蛋白抗原建立的间接ELISA的检测符合率为88.9%。用该方法对黑龙江省、吉林省、河北省、上海市、辽宁省等地猪场送检的257份血清样品进行检测,PCV1和PCV2血清抗体阳性检出率分别为19.1%和80.5%,PCV1抗体阳性猪中有95.9%为PCV2阳性,表明我国猪群已存在PCV1感染,在临床上PCV1与PCV2多以混合感染形式存在。建立的PCV1-IPMA方法具有操作简便、特异和敏感等特点,不失为PCV1流行病学调查及血清抗体检测的有效技术手段。

An immunoperoxidase monolayer assay（IPMA） was developed for detection of antibodies against porcine cireovirus type 1 （PCV1）. The reaction conditions, specificity, sensitivity, and repeatability of IPMA were optimized, evaluated, and compared with those of ELISA based on the baculovirus-expressed PCV1 recombinant capsid protein as coating antigen. Different dilutions of PCV1 and PCV2 positive reference sera were assayed by IPMA,and the low level of cross-reaction of antibody against PCV1 with that against PCV2 could be eliminated by serum dilution of more than 1 ： 100. There was no cross-reaction with reference sera against other porcine viruses. In comparison,88.9% agreement rate was obtained between IPMA and ELISA. A total of 257 serum samples from Heilongjiang, Jilin, Hebei, Shanghai, Liaoning provinces were detected using PCVI-IPMA and PCV2qPMA. The positive rate was 19.1 % and 80.5 % for PCV1 and PCV2, respectively. There were antibodies against PCV2 in approximately 95. 9% of PCV1- positive samples, indicating that mixed infectious of PCV1 with PCV2 existed in the above-mentioned provinces. The results of comparison showed that the developed PCVI-IPMA offered a convenient tool for the epidemiological investigation of PCV1 infections and evaluation of antibodies against the virus.