摘要
本文探讨了用JEV—E McAb制备亲和层析柱纯化E糖蛋白的方法。结果表明,超声粉碎法裂解病毒囊膜不影响HA活性,而用NP40、去氧胆酸钠、Tri—ton X-100处理病毒原始材料,则导致HA活性的丧失;pH11.5、0.05M二乙胺洗脱剂对E的生物活性影响较少,而3M KSCN对E的HA活性有明显影响;用亲和力适中的McAb2F_2和mC_3亲和层析柱纯化E糖蛋白,其HA活性的回收率分别为30～40%及30～60%,蛋白收获量分别为0.33～1.26mg及0.21～0.66mg,相对HA比活性分别提高3～8倍及8～30倍;经SDS—PAGE考马斯亮兰染色,显示一条带,分子量相当于E糖蛋白;纯化的E糖蛋白可诱导小鼠产生HI抗体及NT抗体。
This paper reported a purification method to isolate JEV-E glycopro-tein in mg amount using McAb affinity chromatography. The results indi-cated that the HA activity of the viral primary material was not affected byultrasonic treatment but destroyed in splitting with NP40, deoxycholate, or tri-ton X-100. The biologic activity of E eluated by 0.05M diethylamine wasstill retained, but lost when 3M KSCN was used. The 2F_2 and mC_3 McAbwhich were moderate in affinity was used in immunoadsorption chroma-tography to purify JEV-E glycoprotein from crude JEV suspention. Theyeild of HA actiyity of E was 30～40% and 30～60%, and the proteincontent was 0.33～1.26mg and 0.21～0.66mg, so that the relative specificactivity increased 3～8 times and 8～30 times respectively. SDS～PAGEshowed that tbe molecular weight of the two eluates are identical (52.3kd). Mice immunized with purified E developed HI and NT antibodies.
出处
《解放军预防医学杂志》
CAS
北大核心
1992年第1期23-27,共5页
Journal of Preventive Medicine of Chinese People's Liberation Army
基金
中国科学院科学基金
关键词
日本脑炎病毒
E糖蛋白
单克隆抗体
Japanese encephalitis virus (JEV)
E-glycoprotein
monoclonal antibodies (McAb)
affinity chromatography
