Transfection of enhanced green fluorescent protein into immortalized neural progenitor cells by nonª²viral vectors

    GUI Lingª²Li, LIU Zhiª²Heng, ZHANG Chuanª²Han, GAO Feng

    1Department of Anesthesiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China

    2Department of Anesthesiology, Second People¬ðs Hospital of Shenzhen City, Shenzhen 518035, China

    ¡¾Abstract¡¿ AIM: To investigate an effective transfection method of nonª²viral vectors and the expression of enhanced green fluorescent protein (EGFP) in the transfected immortalized neural progenitor cells (INPC). METHODS: The plasmid EGFPª²C1 was transfected into INPC by three different nonª²viral vectors respectively, including Lipofectamine 2000, TRANSfection and Sofast. The expression of EGFP and the transfection efficiency of the 3 vectors were measured at 24 h after transfection. The transfection efficiency of the vector which was the most efficient, was detected 12, 24, 48 and 72 h after transfection to determine the expression peak of EGFP. The viability of transfected and nonª²transfected cells was measured by trypan blue rejection test. After the plasmid EGFPª²C1 was transfected into INPC, the stable cell clone designated INPC/EGFP was isolated by G418 selection. The positive rate of EGFP was observed. And the specific molecular marker of neural progenitor cells, nestin, was detected using immunocytochemistry. After INPC/EGFP was induced by 50 mL/L fetal bovine serum, the cell morphology and the expression of EGFP were observed in the differentiated cells. RESULTS: The transfection efficiency of Lipofectamine 2000, TRANSfection and Sofast was (25.5¡À2.9)%, (4.0¡À1.7)%, (7.9¡À1.4)% respectively, at 24 h after transfection. And Lipofectamine 2000 was the most efficient. Its transfection efficiency at 12, 24, 48 and 72 h after transfection was (17.1¡À0.7)%, (25.5¡À2.9)%, (19.4¡À0.9)%, (15.6¡À1.4)%, respectively. The expression of EGFP was the highest at 24 h after transfection. The positive rate of EGFP was 95% in INPC/EGFP. And INPC/EGFP was still nestin positive. The differentiated cells presented as neurons or astrocytes, and EGFP was still found in their somas and processes. CONCLUSION: Extrinsic genes can be effectively transfected into INPC by Lipofectamine 2000. And EGFP is one of the valuable tracking tools for INPC. ......