Cationic liposomal delivery of plasmid to endothelial cells measured by quantitative flow cytometry

✍ Scribed by Wenchi Tseng; Norman B. Purvis; Frederick R. Haselton; Todd D. Giorgio

• Publisher: John Wiley and Sons
• Year: 2000
• Tongue: English
• Weight: 758 KB
• Volume: 50
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/(sici)1097-0290(19960605)50:5<548::aid-bit9>3.0.co;2-f

✦ Synopsis

Cationic liposornes are potentially important gene transfer vehicles, capable of conjugating with anionic DNA by condensation. Flow cytometry was used to examine quantitatively the incorporation of DNA-liposome complex into murine capillary lung endothelial cells. The plasmid DNA, a pSV-p-galactosidase vector, was covalently labeled with ethidiurn monoazide by photoactivation. The cationic liposome consisted of egg phosphatidylcholine (go%), cholesterol (5%), and stearylamine (5%). The number of plasmid molecules contained within each cell as a function of exposure time was estimated from fluorescence intensity. Fluorescently labeled plasmid is detectable after 10 min and increases with continued exposure, but at a decreasing rate, up t o 2160 min. After 2160 min each cell, on average, contains approximately 10,000 plasmid molecules. Following transfection, a single cell unirnodal population was detected by flow cytometry, suggesting that all cells participate in transfection equally. Furthermore, cell cycle analysis indicates that the entry of DNA-liposome complex is independent of cell cycle.