华支睾吸虫成虫RPEF基因产物及其抗RPEF多克隆抗体的制备

The preparation of RPEF gene products of Clonorchis sinensis and the anti-RPEF polyclonal antibodies

目的体外制备华支睾吸虫成虫RNA聚合酶Ⅱ延长因子（RPEF）基因产物及抗RPEF多克隆抗体。方法亲和纯化试剂盒纯化表达蛋白pGEx4T-1，RPEF，凝血酶酶切表达蛋白制备重组蛋白RPEF，将重组蛋白RPEF免疫小鼠，制备多克隆抗血清，检测抗体滴度和抗血清特异性。结果体外制备的RPEF蛋白经SDS电泳呈单一条带；酶联免疫吸附结果显示，免疫过程抗体滴度呈连续上升趋势；免疫印迹结果显示，小鼠抗血清具有抗RPEF特异性。结论获得较专一的华支睾吸虫RPEF蛋白和RPEF抗血清。

Objective To prepare RPEF Gene products of Clonorchis sinensis and the anti-RPEF polyclonal antibodies. Methods The expressed protein pGEX-4T-1-RPEF was purified by GST Bind Purification kit and was digested with thrombin. The experimental mice were inoculated with recombinant RPEF protein. The polyclonal antisera were obtained from the immuned mice. The antisera collected were used to detect the titres of IgG antibodies and to identify anti-RPEF specificity. Results SDS-PAGE showed single band of the purified RPEF protein. ELISA analysis showed that IgG titer in the immuned mice was increased. The results of West- ern blot suggested that the mouse antir RPEF antisera were highly specific. Conclusion The RPEF gene products of Clonorchis sinensis and the anti-RPEF polyclonal antibodies were successfully prepared.