鸭乙型肝炎病毒表面抗原的纯化及应用

Preparation and identification of duck hepatitis B virus (DHBV) surface antigen from infected duck serum

目的：从鸭乙型肝炎病毒（DHBV）感染鸭血清中纯化表面抗原（DHBsAg），初步应用于病毒感染后复制的研究。 方法：用蔗糖密度梯度离心纯化病毒表面抗原，经Bradford法定量血清中该蛋白含量； 建立DHBsAg ELISA检测方法， 并与DHBV DNA斑点杂交相比较。 结果：纯化DHBsAg 经ELISA检测OD490nm≥1.0，Western杂交显示主要肽分子量为17KDa和35KDa；Bradford法定量血清蛋白含量为2.85～5.10μg/ml, DHBsAg ELISA检测100份血清标本阳性数为84例, 斑点杂交检测阳性数为88例,两者吻合率为94%。 结论：DHBsAg 的制备及ELISA方法的建立，为进一步研究病毒感染后复制及体内免疫应答状况奠定了基础。

Objective: Preparation of DHBsAg from DHBV infected ducks and its application for the study of virus replication. Methods : The positive serum were layered over a step gradient of 20% and 66% sucrose, DHBsAg were harvested at the interface between 20% and 66% sucrose, and then analyzed by Bradford method. Meanwhile, DHBsAg ELISA and DHBV DNA dot blot hybridization assayed 100 serum samples. Results : Purified antigen were detected OD490nm≥1.0 by DHBsAg ELISA assay and quantified 2.85~5.10μg/ml by Bradford method. Both 35KDa and 17KDa pres/s bands were present in DHBV-infected serum. DHBsAg assayed 84 positive in 100 serum samples, while DHBV DNA dot blot hybridization detected 88 positive, showing good accordance. Conclusion: Preparation of DHBsAg from DHBV-infected ducks and application of DHBsAg ELISA assay will be helpful to the study of kinetics of DHBV and immune response of ducks to the virus.