摘要
为建立快速检测猪流感病毒抗体的血清学方法,利用合成的甲型流感病毒M2蛋白多肽作为包被抗原,建立了检测猪流感病毒血清抗体的间接ELISA,并对各种检测条件进行了优化。结果显示,优化后确定的抗原最适包被量为250ng/孔,血清最佳稀释度为1∶100,酶标二抗最佳稀释度为1∶4 000。在优化条件下,阴阳性临界值的判定标准为0.249。用建立的间接ELISA对CSFV、PRRSV、PCV2、PPV、PRV、FMDV、JEV阳性血清进行了检测,均无交叉反应。用该方法对280份冬季采集的猪血清样品进行检测,有60份样品呈现阳性;同时用IDEXX甲型流感病毒抗体检测试剂盒进行平行检测,有61份样品呈现阳性,两者的符合率达到98.35%。结果表明,建立的ELISA具有良好的敏感性、特异性和可信度。本研究为猪流感的诊断和流行病学调查提供了一种准确、快速、简便的方法。
An indirect ELISA was established for rapid detection of antibodies against swine influenza virus（SIV） using synthetic M2 protein polypeptide of influenza A virus as coating antigen. The reaction conditions were optimized,including 250 ng/well coating antigen of synthetic SIV M2 protein polypeptide, 1 ： 100 dilution of testing serum and 1 ： 4 000 dilution of HRP-conjugated Rabbit Anti-pig IgG with cut-off value 0. 249（D_450mm）. No cross-reaction was observed with the positive sera against classical swine fever virus, porcine reproductive and respiratory syndrome virus, porcine circovirus, porcine parvovirus, pseudo- rabies virus,foot-and-mouth disease virus and Japanese encephalitis virus,respectively. A total of 280 ser- um samples collected in winter were tested,60 samples were positively detected by the indirect ELISA and 61 samples showed positive detected by IDEXX SIV ELISA,the agreement rate was 98.35%. All these re- sults show that the established ELISA method has high sensitivity, reliability and credibility. This study provides an accurate,rapid and simple method for the diagnosis and epidemiological investigation of SIV.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2015年第7期700-705,共6页
Chinese Veterinary Science
基金
教育部"长江学者和创新团队发展计划"项目(IRT13083)
教育部新世纪优秀人才支持计划项目(NCET11-1059)
