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Assessment of Different Transfection Parameters in Efficiency Optimization

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Abstract:

Achieving optimal transfection efficiency is the most critical step in overcoming the primary obstacle to success in nonviral-mediated gene therapy. Several transfection parameters were being examined including the effects of different types of transfection media, glucose concentration, reporter DNA concentration, and incubation time in lipotransfectant. Efficiency of transfection obtained was highest for Opti-MEM I (29 ± 2.28%; p = 0.001) followed by M199 (24 ± 1.54%; p = 0.009), both of which performed significantly better than DMEM (14 ± 0.28%) as a transfection medium. The rate of transfection was affected by glucose levels in only DMEM with higher efficiency achieved using low glucose containing DMEM (17 ± 0.38%) than its counterpart. Furthermore, transfection rate and cell viability were severely hampered by lengthened exposure to transfection complexes, leading to an overall mean efficiency of 5 ± 0.87%. However, doubling the DNA content in the transfection mixture did not significantly change the mean rate of transfection in M199 medium (24 ± 1.54% to 27 ± 1.54%; p = 0.273). The overall range of mean efficiency acquired with our protocol under different transfection conditions was between 14% and 29%. Hopefully results from this study will further potential success in nonviral-mediated gene transfer.

Keywords: Effectene; Glucose; HUVECs; Transfection efficiency; Transfection media

Document Type: Review Article

DOI: https://doi.org/10.3727/000000004773301861

Affiliations: 1: *Department of Surgery, Surgical-Medical Research Institute,University of Alberta, Edmonton, Canada T6G 2N8 2: †Department of Medicine, University of Alberta, Edmonton, Canada T6G 2N8

Publication date: 2004-01-01

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  • Cell Transplantation publishes original, peer-reviewed research and review articles on the subject of cell transplantation and its application to human diseases. To ensure high-quality contributions from all areas of transplantation, separate section editors and editorial boards have been established. Articles deal with a wide range of topics including physiological, medical, preclinical, tissue engineering, and device-oriented aspects of transplantation of nervous system, endocrine, growth factor-secreting, bone marrow, epithelial, endothelial, and genetically engineered cells, among others. Basic clinical studies and immunological research papers are also featured. To provide complete coverage of this revolutionary field, Cell Transplantation will report on relevant technological advances, and ethical and regulatory considerations of cell transplants. Cell Transplantation is now an Open Access journal starting with volume 18 in 2009, and therefore there will be an inexpensive publication charge, which is dependent on the number of pages, in addition to the charge for color figures. This will allow work to be disseminated to a wider audience and also entitle the corresponding author to a free PDF, as well as prepublication of an unedited version of the manuscript.
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