Raptis L H, Brownell H L, Liu S K, Firth K L, MacKenzie L W, Stiles C D, Alberta J A
Department of Microbiology and Immunology, Queen's University, Kingston, Ontario, Canada.
Mol Biotechnol. 1995 Oct;4(2):129-38. doi: 10.1007/BF02921607.
Nontraumatic, simple, and reproducible procedures for the introduction of nonpermeant molecules into adherent mammalian cells by in situ electroporation are described. Cells are grown on a glass slide, half of which is coated with electrically conductive, optically transparent, indium-tin oxide. An electric pulse is applied in the presence of the molecules to be introduced, and their effect on the cellular phenotype can be observed. The cells growing on the nonconductive side of the slide do not receive any pulse and serve as controls. Careful adjustment of electric field strength can achieve the introduction of the molecules into essentially 100% of the cells, and this treatment causes no detectable disruption to cellular metabolism. This is applied in the presence of the fluorescent dye, Lucifer yellow, causing its penetration into the cells growing on the conductive half of the slide. The migration of the dye to the nonelectroporated cells growing on the nonconductive area is microscopically observed under fluorescence illumination.
本文描述了通过原位电穿孔将非渗透性分子导入贴壁哺乳动物细胞的非创伤性、简单且可重复的方法。细胞生长在载玻片上,载玻片的一半涂有导电、光学透明的氧化铟锡。在待导入分子存在的情况下施加电脉冲,并可观察其对细胞表型的影响。生长在载玻片非导电侧的细胞不接受任何脉冲,作为对照。仔细调整电场强度可使分子基本上100%导入细胞,且这种处理对细胞代谢无明显破坏。在荧光染料路西法黄存在的情况下应用此方法,可使其渗透到生长在载玻片导电半侧的细胞中。在荧光照明下用显微镜观察染料向生长在非导电区域的未电穿孔细胞的迁移情况。