Electroporation by nucleofector is the best nonviral transfection technique in human endothelial and smooth muscle cells
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BioMed Central
Open Access
Methodology
Electroporation by nucleofector is the best nonviral transfection technique in human endothelial and smooth muscle cells Nina Iversen*, Baard Birkenes, Kari Torsdalen and Srdjan Djurovic Address: Department of Medical Genetics, Ullevål University Hospital, Oslo, Norway Email: Nina Iversen* - [email protected]; Baard Birkenes - [email protected]; Kari Torsdalen - [email protected]; Srdjan Djurovic - [email protected] * Corresponding author
Published: 18 April 2005 Genetic Vaccines and Therapy 2005, 3:2
doi:10.1186/1479-0556-3-2
Received: 07 December 2004 Accepted: 18 April 2005
This article is available from: http://www.gvt-journal.com/content/3/1/2 © 2005 Iversen et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
ElectroporationGene TherapyLiposomesLipofectionPhotochemical InternalizationNucleofectionTransfection
Abstract Background : The aim of this study was to determine the optimal non-viral transfection method for use in human smooth muscle cells (SMC) and endothelial cells (EC). Methods: Coronary Artery (CoA) and Aortic (Ao) SMC and EC were transfected with a reporter plasmid, encoding chloramphenicol acetyltransferase type 1 (CAT), with seven different transfection reagents, two electroporation methods and a photochemical internalization (PCI) method. CAT determination provided information regarding transfection efficiency and total protein measurement was used to reflect the toxicity of each method. Results: Electroporation via the nucleofector machine was the most effective method tested. It exhibited a 10 to 20 fold (for SMC and EC, respectively) increase in transfection efficiency in comparison to the lipofection method combined with acceptable toxicity. FuGene 6 and Lipofectamine PLUS were the preferred transfection reagents tested and resulted in 2 to 60 fold higher transfection efficiency in comparison to the PCI which was the least effective method. Conclusion: This study indicates that electroporation via the nucleofector machine is the preferred non-viral method for in vitro transfection of both human aortic and coronary artery SMC and EC. It may be very useful in gene expression studies in the field of vascular biology. Through improved gene transfer, non-viral transfer techniques may also play an increasingly important role in delivering genes to SMC and EC in relevant disease states.
Background Several methods have been described to introduce DNA expression vectors into mammalian cells in vitro and in vivo: calcium phosphate precipitation, microinjection, electroporation, receptor-mediated gene transfer, particle guns, viral vectors, and lipofection [1-3]. Each system has
benefits and limitations, and to date there is no ideal method for gene trans
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