以J-Lat 11.1为HIV-1潜伏感染模型再激活效果检测方法的初步探讨  

Preliminary Study on the Detection Methods of Reactivation Based on HIV-1 Latency Reactivation Model J-Lat 11.1 Cell Line

作　　者：李莉莉[1] 杨怡姝[1] 王小利[1] 

机构地区：[1]北京工业大学生命科学与生物工程学院,北京100124

出　　处：《生物技术通讯》2016年第4期497-500,共4页Letters in Biotechnology

基　　金：国家科技重大专项(2014ZX10005-002);抗病毒药物北京市国际科技合作基地项目;"2011计划"生物治疗协同创新中心资助项目

摘　　要：目的：比较3种检测方法的优缺点,探索全面评价人免疫缺陷病毒1型（HIV-1）潜伏感染再激活剂的检测方法。方法：以HIV-1潜伏细胞株J-Lat 11.1为潜伏感染模型、豆蔻酰佛波醇乙酯（PMA）为潜伏再激活剂,用流式细胞术检测绿色荧光蛋白（GFP）阳性细胞所占比例,酶标仪检测GFP表达强度,活细胞成像系统检测GFP的动态表达情况。结果：流式细胞术检测显示PMA再激活出GFP阳性细胞的比例随作用浓度的增加（0~10 nmol/L）而增加,但当PMA浓度高于10 nmol/L后变化不再明显;酶标仪检测显示PMA处理后24 h内,GFP荧光强度逐渐增高,之后可在高水平保持至48 h;活细胞成像系统则可以动态反映PMA处理后的再激活过程。结论：可采用流式细胞术检测GFP阳性细胞比例对HIV-1潜伏感染再激活剂进行初步筛选,再结合酶标仪或活细胞成像系统动态监测GFP的表达情况,综合评价再激活剂的作用强度和起效时间。Objective： To explore the detection methods for comprehensively evaluating latency reactivationagents, three methods were compared. Methods： J-Lat 11.1 cell line was adopted as HIV-1 latency reactivationmodel, in which PMA worked as positive reactivation agent. The percentage of GFP positive cells, the GFP intensi-ty and the dynamic expression of GFP were detected with flow cytometry, enspire multimode plate reader or Zeisslive cell imaging system respectively. Results： Flow cytometry showed that the proportion of GFP positive cells in-creased with the escalation of the concentration of PMA（0~10 nmol/L）, but the change was no longer significantafter the PMA concentration was higher than 10 nmol/L. Multimode microplate assay showed that the fluorescenceintensity gradually increased within 24 h after PMA treatment, which maintained at a high level till 48 h. Livecell imaging system can reflect the reactivation process dynamically after PMA treatment. Conclusion： To conducta preliminary screening, flow cytometry was applied to detect the proportion of GFP positive cells. Then multimodemicroplate assay or live cell imaging system was recommended to comprehensively evaluate the reactivation effectand onset time.

关 键 词：人免疫缺陷病毒1型 潜伏感染 再激活 J-Lat细胞 

分 类 号：Q25[生物学—细胞生物学] R392.1[医药卫生—免疫学]