小RNA转染人外周血原代T淋巴细胞的转染试剂筛选

发布时间：2018-03-31 21:41

本文选题：RNA干扰　切入点：转染　出处：《中国医药生物技术》2014年05期

【摘要】：目的探讨人工合成RNA转染人外周血原代T淋巴细胞的有效方法。方法从人外周血分离原代T淋巴细胞,采用Lipofectamine2000、Lipofectamine RNAiMAX、FuGENE6、Hiperfect和EntransterTM-R 5种不同转染试剂介导荧光标记的RNA转染原代T淋巴细胞,通过荧光显微镜观察及流式细胞术检测进行转染效率评价。结果分离的人外周血原代T淋巴细胞的纯度达到87.4%。采用上述5种转染试剂介导荧光标记的RNA转染T淋巴细胞,24 h后转染效率分别为(14.81±1.03)%、(14.33±1.24)%、(2.10±0.16)%、(1.69±0.08)%、(72.24±1.26)%。采用EntransterTM-R转染100及50 nmol/L对照RNA,24 h后转染效率分别为73.7%和56.0%;EntransterTM-R转染100 nmol/L对照RNA后48、72 h荧光标记细胞阳性率分别为62.0%、56.6%。结论转染试剂EntransterTM-R可有效介导人工合成RNA转染人外周血原代T淋巴细胞。
[Abstract]:Objective to investigate the effective method of transfection of human primary T lymphocytes with synthetic RNA. Methods Primary T lymphocytes were isolated from human peripheral blood and transfected into primary T lymphocytes by Lipofectamine RNAiMAXFGENE6 HiPerfect and EntransterTM-R. The transfection efficiency was evaluated by fluorescence microscopy and flow cytometry. Results the purity of the primary T lymphocytes isolated from human peripheral blood was 87.4%. The transfection of T lymphocytes was mediated by the five transfection reagents and labeled by RNA. After 24 hours, the transfection efficiency was 14.81 卤1.03 and 14.33 卤1.24 respectively. The positive rates of EntransterTM-R transfection reagent EntransterTM-R were 73.7% and 56.0%, respectively. The positive rates of fluorescent labeled cells were 62.0% and 56.72 h after transfection of 100 nmol/L control RNA and 50 nmol/L TM-R, respectively. Conclusion the positive rates of EntransterTM-R transfection reagent can be 62.0% and 56.6 h after transfection of 100 nmol/L control RNA and 50 nmol/L TM-R, respectively, and the transfection efficiency is 73.7% and 56.0%, respectively. Conclusion the positive rates of EntransterTM-R transfection reagent EntransterTM-R can be 62.0% and 56.6 h after transfection of 100 nmol/L control RNA and 50 nmol/L TM-R, respectively, and the transfection efficiency is 14.81 卤1.03 and 14.33 卤1.24 and 2.10 卤0.16, respectively. Transfection of human peripheral blood primary T lymphocytes by artificial synthesis of RNA.
【作者单位】：首都医科大学附属北京友谊医院肝病中心;首都医科大学附属北京友谊医院肝硬化转化医学北京市重点实验室;首都医科大学附属北京友谊医院普外科;
【基金】：国家自然科学基金(81000172、81370544) 北京市自然科学基金(7142043) 王宝恩肝纤维化研究基金(20120131)
【分类号】：R392

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