原位RT-PCR法定位检测柯萨奇B病毒RNA

Experimental Study on Detection of Viral RNA of Coxsackie Virus B(CBV)by Means of in Situ RT-PCR

目的 用定位检测细胞或组织中柯萨奇B病RNA的原位RT PCR方法 (直接法 )。方法 对柯萨奇B病毒 ( 1～ 6型 )感染HeLa细胞 ,经核酸酶 (DNase和RNase)处理后的病毒感染HeLa细胞以及非感染的HeLa细胞进行RT PCR和原位RT RCR检测。结果 经原位RT PCR扩增后 ,病毒感染的HeLa细胞呈蓝黑阳性信号 ;并且这种阳性信号经RNaseA作用而消失 ,却不受DNase作用的影响 ;而非感染的HeLa细胞则不着色 ,其显示的结果与RT PCR的结果一致。结论 原位RT PCR法可特异地检测柯萨奇B病毒感染HeLa细胞中的病毒RNA ,是一种特异和敏感的定位检测方法 ,可应用于研究该病毒的持续性感染及其与克山病等相关疾病的关系。

Objective To detect viral RNA in Coxsackie virus B infecting HeLa cells.Method No-viral infecting HeLa cells and coxsackie virus B infecting HeLa cells are digested with nucleases by means of in situ RT-PCR.Results No-virally infected HeLa cells are negative while virally infected HeLa cells are stained with blue and black color,and the positive signals could be removed after digestion with RNase but not DNase.Conclusion The method of in situ RT-PCR is specific and sensitive,and will be useful for the research of Coxsackie virus B infection and related diseases.