CAJ | 학술논문

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 Lipofectamine 2000、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。
목적탐토인공합성 RNA전염인외주혈원대 T림파세포적유효방법。방법종인외주혈분리원대 T림파세포，채용 Lipofectamine 2000、Lipofectamine RNAiMAX、FuGENE6、Hiperfect화 EntransterTM-R 5충불동전염시제개도형광표기적 RNA전염원대 T림파세포，통과형광현미경관찰급류식세포술검측진행전염효솔평개。결과분리적인외주혈원대 T림파세포적순도체도87.4%。채용상술5충전염시제개도형광표기적 RNA전염 T림파세포，24 h후전염효솔분별위（14.81±1.03）%、（14.33±1.24）%、（2.10±0.16）%、（1.69±0.08）%、（72.24±1.26）%。채용 EntransterTM-R전염100급50 nmol/L대조 RNA，24 h후전염효솔분별위73.7%화56.0%；EntransterTM-R전염100 nmol/L대조 RNA후48、72 h형광표기세포양성솔분별위62.0%、56.6%。결론전염시제 EntransterTM-R가유효개도인공합성 RNA전염인외주혈원대 T림파세포。
Objective To investigate the efficient way by which synthetic RNA could be transfected into human primary T-lymphocytes of peripheral blood. Method Primary T-lymphocytes were separated from human peripheral blood. Isolated T-lymphocytes were transfected with fluorescently labeled RNA by different transfection reagents including Lipofectamine 2000, Lipofectamine RNAiMAX, FuGENE6, Hiperfect and EntransterTM-R. Transfection efficiency of each transfection reagent was assessed by fluorescence microscopy and further determined by flow cytometry. Result The purity of isolated T-lymphoctyes was 87.4%. Twenty four hours after fluorescently labeled RNA tranfected T lymphocytes by Lipofectamine 2000, Lipofectamine RNAiMAX, FuGENE6, Hiperfect and EntransterTM-R, transfection efficiency was (14.81 ± 1.03)%, (14.33 ± 1.24)%, (2.10 ± 0.16)%, (1.69 ± 0.08)%, (72.24 ± 1.26)%, respectively. When fluorescently labeled RNA of 100 and 50 nmol/L tranfectd T-lymphocytes by EntransterTM-R, transfection efficiency detected by flow cytometry 24 hours later were 73.7% and 56.0%,respectively. Forty eight hours and seventy two hours after 100 nmol/L fluorescently labeled RNA transfection by EntransterTM-R, the percentage of fluorescence positive cells detected by flow cytometry were 62.0% and 56.6%, respectively. Conclusion Synthetic RNA could be efficiently transfected into human primary T lymphocytes of peripheral blood by EntransterTM-R.