pH dependent high transfection efficiency of mouse neuroblastomas using TransFectin

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• 作者：Kevin Wai Yin Chong ; Alan Yiu-Wah Lee ; Evelyn S.C. Koay ; Sze Jee Seet and Nam Sang Cheung
• 关键词：pH ; Mouse ; Neuroblastoma ; Transfection ; TransFectin™ ; Neuro-2A ; N1E-115 ; Optimised procedure
• 刊名：Journal of Neuroscience Methods
• 出版年：2006
• 出版时间：15 November 2006
• 年：2006
• 卷：158
• 期：1
• 页码：56-63
• 全文大小：947 K

文摘

Mouse neuroblastoma cell lines are often used in lieu of mouse primary neurons in ex vivo experiments, as they provide an easier platform for transfection, compared to the latter. A well-known inherent problem with this strategy is the relatively low transfection efficiency (15–30 % ) of mouse neuroblastoma cell lines such as neuro-2A and N1E-115. We were able to improve the transfection efficiency of these cell lines by using the cationic lipid reagent, TransFectin™ (Bio-Rad, Hercules, CA, USA) to optimise the transfection conditions. Our results, based on fluorescence intensity determinations and Western blotting for enhanced green fluorescence protein (EGFP) over-expression in neuro-2A, demonstrated that pH is a crucial factor in determining the transfection efficiency. Under pH-optimised transfection conditions, flow cytometric analysis revealed high EGFP transfection efficiencies of 76.4 ± 0.5 and 60.9 ± 0.6 % for neuro-2A and N1E-115, respectively. Notably, the optimised TransFectin™-based transfection system did not result in any detectable cytotoxicity to the mouse neuroblastomas. The resultant optimised system is economical, easy to use and does not require any specialised equipment.