猪伪狂犬病病毒和猪繁殖与呼吸综合征病毒抗体同步检测的液相芯片技术

摘要

为了建立猪伪狂犬病毒抗体和猪繁殖与呼吸障碍症病毒抗体的同步检测技术,通过蛋白抗原偶联微球,建立单重和双重液相芯片检测方法,并对蛋白抗原偶联微球量进行优化,验证该方法的特异性,同时用此方法与ELISA试剂盒血清检测方法进行灵敏度比较.结果证明,猪繁殖与呼吸综合征病毒 N 蛋白和猪伪狂犬病毒GE蛋白的抗原最佳偶联量分别为每2.5×105个微球偶联8 μg和2.5 μg;特异性试验结果显示,这两个蛋白抗原偶联的微球与猪其他病原的抗体无交叉反应,说明特异性良好;与 ELISA 方法比较发现,针对同份猪血清样品,液相芯片法的猪伪狂犬病毒和猪繁殖与呼吸综合征病毒抗体的最高血清检出稀释倍数比ELISA法分别高66倍和32倍.说明成功建立了一种同步检测猪伪狂犬病毒和猪繁殖与呼吸综合征病毒抗体的液相芯片方法.%To establish a serological method for the simultaneous detection of porcine pseudorabies virus antibody and porcine reproductive and respiratory disease virus antibody and to provide new technical sup-port for the simultaneous detection of these two swine virus antibodies,antigens were couplied with micro-spheres for establishing single and double liquid chip detection methods,and optimizing the amount of pro-tein antigen coupling microspheres,verifying the specificity of the method.The serum detection sensitivity of method was compared with ELISA kit.The results showed that the optimum coupling amounts of anti-gen of porcine reproductive and respiratory disease virus N protein and PRV GE protein were 8 μg and 2.5 μg per 2.5×105microspheres respectively.The result of specific experiment showed that two protein anti-gen-conj ugated microspheres has no cross-reactivity with other pig pathogen antibodies,indicating good specificity;compared with the ELISA method,the result showed that for the same pig serum samples,the limits of serum dilutions detected with antibodies were 32 and 66 fold higher respectively.In conclusion, this study successfully established a method for simultaneous detection of antibodies of porcine pseudora-bies virus and porcine reproductive and respiratory disease virus.