Gene delivery process in a single animal cell after femtosecond laser microinjection
Gene delivery process in a single animal cell after femtosecond laser microinjection
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DOI：
10.1016/j.apsusc.2009.04.111
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发表时间：
2009-09-30
影响因子：
6.7
通讯作者：
Masuhara, Hiroshi
中科院分区：
文献类型：
--
作者：
Hosokawa, Yoichiroh;Iguchi, Seriya;Masuhara, Hiroshi
通过放大的飞秒激光照射将细胞外分子显微注射到单个动物细胞中。当用高数值孔径物镜透镜将单次激光脉冲聚焦在来自小鼠细胞系NIH 3 T3的单个成纤维细胞的质膜上时，形成直径为1 mm的瞬时孔。通过用荧光素异硫氰酸盐-葡聚糖（FITC-葡聚糖）进行荧光染色来监测孔形成后细胞外分子的递送过程。然后使用增强型绿色荧光蛋白（EGFP）的DNA质粒确认基因表达。当激光脉冲首先聚焦在细胞膜上，然后聚焦在核膜上时，观察到基因表达，而当激光仅聚焦在细胞膜上时，基因不表达。在这些结果的基础上，阐明了飞秒激光显微注射的基因递送效率和随后的基因表达。(C)2009 Elsevier B. V.保留所有权利。
Microinjection of extracellular molecules into a single animal cell was performed by an amplified femtosecond laser irradiation. When a single-shot laser pulse was focused on the plasma membrane of a single fibroblast from the mouse cell line NIH3T3 with a high-numerical aperture objective lens, a transient hole with a diameter of 1 mm was formed. The delivery process of extracellular molecules immediately after the hole formation was monitored by a fluorescence staining with fluoresceini-sothiocyanate-dextran (FITC-dextran). Then the gene expression was confirmed using a DNA plasmid of an enhanced green fluorescent protein (EGFP). The gene expression was observed when the laser pulse was focused first on the cellular membrane and then on the nuclear membrane, while the gene was not expressed when the laser was focused only on the cellular membrane. On the basis of these results, the efficiency of gene delivery by the femtosecond laser microinjection and the subsequent gene expression were clarified. (C) 2009 Elsevier B.V. All rights reserved.