猫肾F_(81)传代细胞中犬细小病毒原位PCR检测方法

Study of in situ Polymerase Chain Reaction Assay for Detection of Canine Parvovirus(CPV) in Feline Kidney F_(81) Cells

目的 建立在猫肾F81 细胞中犬细小病毒原位PCR的检测方法。方法 在猫肾F81 细胞上感染犬细小病毒 ,设计特异性引物 ,用直接原位PCR法在染毒 12h ,2 4h ,48h细胞片上检测出犬细小病毒 ,并与常规免疫组化的方法进行了比较。结果 在染毒 48h的细胞片上 ,用阳性记分法将两种检测方法得到的阳性细胞进行统计比较 ,差异极显著 (P <0 .0 0 1) ,用原位PCR法所得出的阳性率高。

Objective To Study of in situ polymerase chain reaction assay for the detection of canine parvovirus(CPV) in Feline Kidney F 81 cells.Method Feline kidney F 81 cells were cultured in slide and infected with CPV. The cells infected with CPV in different stages were detected by direct in situ PCR, which used biotin labelled primers, and immunohistochemical method. Result In situ PCR method, a lot of signals were found in cells infected from 12 to 48 hours. Some signals were found in cytoplasm, and some in nucleus. In immunohistochemical method, no signals were found in cells infected for 12 hours, and sporadic signals in cells infected for 48 hours. The signals detected by these two methods showed significant difference. Conclusion The results indicate that the in situ PCR not only has high sensitivity, but also can locate the position of virus.