甲型肝炎病毒在3株细胞中的增殖和连续传代  

Propagation and continuous passage of HAV in three cell lines

作　　者：郑纪山[1] 何亮[1] 王宏[2] 周宗安[1] 邓小昭[1] 乔仁良[1] 

机构地区：[1]南京军区联勤部军事医学研究所分子生物学研究室,江苏南京210002 [2]东南大学分子与生物分子电子学实验室,江苏南京210096

出　　处：《医学研究生学报》2001年第4期294-296,299,共4页Journal of Medical Postgraduates

摘　　要：目的 :观察甲型肝炎病毒 (HAV )在 3个不同的细胞株中连续传代时的增殖特点。　方法 :将 HAV NJ- 3株先以常规传代方式适应不同的细胞株 :FRh K4细胞、PL C/ PRF/ 5细胞、2 BS,然后将适应病毒接种于细胞 ,并随细胞传代。用 EIA、IFA和激光扫描共聚焦显微镜观察带毒细胞连续传代中的病毒增殖。　结果 :1NJ- 3株 / K4细胞于第 5代 HAAg出现阳性 ,第 17、18代时达到高峰 ,2 4代直至 35代均为阴性。 2 NJ- 3株 / 2 BS细胞仅在第 6代出现一次弱阳性 ,传至 35代均呈阴性。 3NJ- 3株 / PL C则在第 3代即可测得 HAAg,第 7代 A值达 0 .80 2 ,P/ N值13.9,并保持此水平至少 70代以上。病毒产量稳定 ,细胞形态良好。冻存 10年的 HAV/ PL C株复苏后 ,仍保持良好的产毒性能。HAV/ PL C传代或收获的最佳周期为 5天。　结论 :HAV NJ- 3株能适应三种不同的传代细胞株 ,但只能在 PL C/ PRF/ 5细胞中长期随细胞传代 。Objectives: To observe the characteristic of propagation and continuous passage of HAV in vitro companying with passing of different cell lines. Methods:The strain of NJ 3 HAV adapted to growth in PLC/PRF/5, FRhK4 and 2BS cell lines were inoculated in these cells, and passed in company with the cells passage. Then HAAg in cells was detected by IFA and LSCM, and the antigen titre was detected by EIA during each passage. Results: ①HAAg produced by HAV/K4 cells could be detected in passage 5, the titre of HAV came to a peak in passage 17 and 18, but it was not detected from passage 24 to 35. ②HAAg produced by HAV/2BS cells was always negative up to passage 35 except weak positive once in passage 6. ③ HAAg produced by HAV/PLC cells could be detected in passage 3.The OD value at 450nm was up to 0.802 and the P/N ratio was 13.9 in passage 7. HAAg production was stable more than 70 passages and PLC cells infected by HAV growthed well without cytopathic effect. The interval of passage or havest of HAV/PLC cells was five days. Conclusions: Strain of NJ 3 HAV could be propagated and continuously passedin PLC/PRF/5 cells in vitro , but could not be in K4 and 2BS cell lines.

关 键 词：甲型肝炎病毒 细胞培养 连续传代 激光扫描共聚焦显微术 

分 类 号：R512.6[医药卫生—内科学]