Surendra Nimesh, Marc Thibault, Marc Lavertu et Michael D. Buschmann
Article de revue (2010)
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Abstract
This study was designed to systematically evaluate the influence of pH and serum on the transfection process of chitosan-DNA complexes, with the objective of maximizing their efficiency. The hydrodynamic diameter of the complexes, measured by dynamic light scattering (DLS), was found to increase with salt and pH from 243 nm in water to 1244 nm in PBS at pH 7.4 and aggregation in presence of 10% serum. The cellular uptake of complexes into HEK 293 cells assessed by flow cytometry and confocal fluorescent imaging was found to increase at lower pH and serum. Based on these data, new methodology were tested and high levels of transfection (>40%) were achieved when transfection was initiated at pH 6.5 with 10% serum for 8-24 h to maximize uptake and then the media was changed to pH 7.4 with 10% serum for an additional 24-40 h period. Cytotoxicity of chitosan/DNA complexes was also considerably lower than Lipofectamine. Our study demonstrates that the evaluation of the influence of important parameters in the methodology of transfection enables the understanding of crucial physicochemical and biological mechanisms which allows for the design of methodologies maximising transgene expression.
Mots clés
Animals; Cattle; *Chitosan; DNA/*administration & dosage/*metabolism; *Gene Transfer Techniques; HEK293 Cells; Humans; Hydrogen-Ion Concentration; Luciferases/analysis; Transfection/*methods
Département: | Département de génie chimique |
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URL de PolyPublie: | https://publications.polymtl.ca/5060/ |
Titre de la revue: | Molecular Biotechnology (vol. 46, no 2) |
Maison d'édition: | Springer |
DOI: | 10.1007/s12033-010-9286-1 |
URL officielle: | https://doi.org/10.1007/s12033-010-9286-1 |
Date du dépôt: | 05 avr. 2022 14:46 |
Dernière modification: | 06 avr. 2024 10:44 |
Citer en APA 7: | Nimesh, S., Thibault, M., Lavertu, M., & Buschmann, M. D. (2010). Enhanced gene delivery mediated by low molecular weight chitosan/DNA complexes: effect of pH and serum. Molecular Biotechnology, 46(2), 182-96. https://doi.org/10.1007/s12033-010-9286-1 |
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