原核和真核细胞表达HBeAg的应用研究

Application of HBeAg produced in prokaryotic cells and eukaryotic cells

目的 :将原核细胞和真核细胞分别克隆表达生产的 HBe Ag,经适当纯化后进行检测分析 ,并在乙型肝炎抗HBe检测中进行应用和比较。 方法 :分别用大肠杆菌和家蚕细胞表达生产 HBe Ag,并用 Saphacryl S- 2 0 0柱层析进行纯化 ;紫外分光光度法测定表达产物的蛋白含量 ;EIA法测定 HBe Ag和 HBc Ag效价及评估 HBe Ag的应用效果。 结果 :原核细胞 HBe Ag:比活性为 10 0 0 0 / m g,HBe Ag/ HBc Ag=5 0 ,用于抗 HBe的检测时特异性为 96 % ,灵敏度符合国家卫生部 panel要求。真核细胞 HBe Ag:比活性为 16 0 0 0 0 / m g,HBe Ag / HBc Ag =5 0 0 0 ,用于抗 HBe的检测时特异性为 10 0 % ,灵敏度高于国家卫生部 panel要求的 1～ 2个滴度。 结论 :真核细胞表达的 HBe Ag比活性高而 HBc Ag含量低 ,在抗

Objectives: To express HBeAg in prokaryotic and eukaryotic cells and compare the two types of HBeAg in the anti HBeAg testing. Methods: HBeAg was expressed both in E.coli cells and in silk worm cells, purified by Sephacryl S 200.HBeAg protein concentration and antigenic titer were determined respectively by ultraviolet spectroscopy and EIA. Results: HBeAg produced by E.coli cells: Activation ratio was 10 000/mg, HBeAg/HBcAg = 50; The specificity in testing anti HbeAg was 96%;HBeAg produced by silk worm cells: Activation ratio was 160 000/mg, HBeAg/HBcAg = 5 000, The specificity in testing anti HbeAg was 100%. Conclusions: HBeAg produced by eukaryotic cells contained much lower proportion of HbcAg and higher activation ratio, which therefore bring about a possibility to improve the quality of the kit for testing Anti HBe.