重组TGFβ_1基因转染后细胞上清的检测

Testing of Cell Culture Fluid by Recombinant TGFβ_1 Gene Transfer

目的 :观察TGFβ1基因转入能否成功。方法 :用pL(TGFβ1)SN经脂质体转染包装细胞 ,G418筛选 ,对阳性病毒上清检测TGFβ1表达蛋白及Neo基因 ,转染后的SMMC 772 1细胞进行免疫组化染色。结果 :阳性上清ELISA检测 ,TGFβ1含量为 16～ 41ng/ml;套式RT -PCR检测Neo基因 ,第 2轮后全部出现 2 90bp条带 ;TGFβ1免疫组化染色显示 :空白对照± ,正义转染 ++。结论 :TGFβ1基因转染效果确实可靠。

Objective: To investigate whether transforming growth factor β 1 gene can transfer successfully or not. Methods: pL( TGF β 1) SN was used to transfect packaging cell line by lipofectamine. After G418 screening, the positive serum of retroviral was detected for TGFβ 1 and Neo gene. Positive SMMC 7721 cell slide for immunohistochemical staining was prepared. Results: The TGF β 1 concentration of cell culture fluid ranged from 16 to 41 ng/ml by ELISA. 290 bp Neo gene band was discovered by nest primer RT-PCR in all the samples. Immunohistochemical staining showed TGF β 1 expression was increased significantly after pL(TGF β 1) SN transfection. Conclusion: The effect of TGF β 1 gene transfer is certain.