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Proceedings Paper

Laser-induced stress wave-assisted gene transfection: improved transfection efficiency with cationic liposome-modified plasmid DNA
Author(s): Risa Otsuka; Mitsuhiro Terakawa; Shunichi Sato; Yasushi Satoh; Kunio Takishima; Hiroshi Ashida; Minoru Obara
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Paper Abstract

Although various methods for gene transfer have been investigated, a practical gene delivery system that fulfills the requirements for clinical application has not yet been developed. Gene transfer by the use of laser-induced stress waves (LISWs) is a physical method to facilitate gene transfer into cells with the effect of stress waves generated by irradiation an absorbing material with high-intensity laser pulses. This method has high spatial controllability and a potential for catheter-based gene transfer. We demonstrated selective high transfection efficiency in vivo. However, there remains a problem that transfection efficiency is limited to less than several percent in vitro. Thus we attempted to improve transfection efficiency by using plasmid DNA modified with cationic-liposome. Plasmid DNA coding for enhanced green fluorescent protein (EGFP) had been modified with Lipofectamine and it was added to a dish for NIH3T3 cell culture. A black rubber disk was placed on the upper side of the cells; the disk was irradiated with 532 nm, nanosecond laser pulses (spot diameter, 3 mm; fluence, 1.3 J/cm2; number of pulses, 20). 24 hours after application of LISWs, transfection efficiency was evaluated with a fluorescence microscope, where efficiency was defined as the ratio of the number of cells emitting fluorescence to the total number of cells. At a DNA concentration of 7.8 &mgr;g/ml, transfection efficiency with naked plasmid DNA was as low as 0.05%, while it was increased to 23.7% by using plasmid DNA modified with Lipofectamine. Since both of the naked plasmid DNA and cell membranes have negative charge, plasmid DNA concentration around cells should be low. Since DNA-Lipofectamine complexes carry positive charge, density of plasmids existing around cells should be increased, resulting in much improved transfection efficiency.

Paper Details

Date Published: 6 February 2007
PDF: 6 pages
Proc. SPIE 6435, Optical Interactions with Tissue and Cells XVIII, 643518 (6 February 2007); doi: 10.1117/12.701734
Show Author Affiliations
Risa Otsuka, Keio Univ. (Japan)
Mitsuhiro Terakawa, Keio Univ. (Japan)
Shunichi Sato, National Defense Medical College (Japan)
Yasushi Satoh, National Defense Medical College (Japan)
Kunio Takishima, National Defense Medical College (Japan)
Hiroshi Ashida, National Defense Medical College (Japan)
Minoru Obara, Keio Univ. (Japan)

Published in SPIE Proceedings Vol. 6435:
Optical Interactions with Tissue and Cells XVIII
Steven L. Jacques; William P. Roach, Editor(s)

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