轮状病毒P[2]G3株在人二倍体细胞KMB17上的适应性  

Adaptivity of Rotavirus P[2]G3 Strain in Human Diploid Cell Line KMB17

作　　者：张光明[1] 刘馨[1] 张永欣[1] 徐婧雯[1] 洪超[1] 孙茂盛[1] 

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所分子生物室,昆明650118

出　　处：《中国生物制品学杂志》2008年第11期969-971,共3页Chinese Journal of Biologicals

基　　金：国家863计划资助项目(2006AA02A211)

摘　　要：目的探讨轮状病毒P[2]G3株在人二倍体细胞KMB17上的适应性。方法轮状病毒P[2]G3株在猴肾细胞MA104上经蚀斑筛选纯化后,得到的克隆以不同MOI接种至人二倍体细胞KMB17上。用微量滴定法与蚀斑法平行检测病毒滴度,用免疫荧光法检测病毒的增殖情况,最后进行病毒基因组稳定性检测。结果经MA104细胞培养的轮状病毒P[2]G3株在人二倍体细胞KMB17上可产生明显的细胞病变(CPE),但病变出现的时间随病毒传代次数的增加而延迟,抗原性也逐渐减弱。通过蚀斑筛选纯化得到的克隆病毒,可在KMB17细胞上稳定地增殖。以0.05MOI病毒量接种可产生稳定的CPE,时间为72～96h。连续传代至第10代,病毒滴度达105.5PFU/ml,病毒基因组稳定。结论轮状病毒P[2]G3株在人二倍体细胞KMB17上表现出毒力及增殖能力的衰退;经蚀斑筛选纯化可获得毒力较强的病毒株,并在KMB17细胞中稳定复制、增殖。Objective To explore the adaptivity of rotavirus P[2]G3 strain in human diploid cell line KMB17. Methods The clones of rotavirus P[ 2]G3 strain cultured in monkey kidney MA104 ceils were purified by plaque screening and inoculated to human diploid cell line KMB17 at various MOI. Determine the virus titer by microtitration and plaque formation test and the virus proliferation by IFA, and perform genomic stability test. Results Obvious CPE was observed in KMB17 cells inoculated with rotavirus P[2]G3 strain after culture in MA104 cells. The antigenieity of virus decreased with the increasing passage, while the appearance of CPE was delayed. The purified P[2]G3 strain was proliferated stably in KMB17 cells. Stable CPE was observed in the KMB17 cells 72-96 h after inoculation with P[2]G3 strain at MOI of 0. 05. The P[2]G3 strain after subeulture in KMB17 cells for 10 passages reached a liter of 10^5.5 PFU/ml and showed good genomic stability. Conclusion Both virulence and proliferation level of rotavirus P[2]G3 strain cultured in MA104 cells decreased after subculture in KMB17 cells. Virulent P[2]G3 strain was purified by plaque screening, and replicated and proliferated stably in KMB17 cells.

关 键 词：轮状病毒 人二倍体细胞 蚀斑筛选 适应性 

分 类 号：R373.9[医药卫生—病原生物学]