Evaluation of different transfection methodologies to achieve efficient expression of the NS1 dengue protein in HepG2 cells

Kíssila Rabelo, Edson R.A. Oliveira, Cecília J. G. Almeida, Ada M.B. Alves, Simone M. Costa

Abstract


HepG2, a human hepatocarcinoma cell line, has been used as a model to study infection by several pathogens including dengue virus. However, this cell line is notoriously difficult to be transfected with plasmid DNAs by traditional methods, which is a limitation for some studies involving heterologous gene expression. In the present work, we analyzed different protocols for transfection of HepG2 with the plasmid pcENS1, which encodes the dengue NS1 protein, in order to evaluate the best methodology for achieving high cell viability and transfection efficiency. We analyzed two transfection approaches using lipid-based methods (Lipofectamine and FuGENE 6) or electroporation by nucleofection. Expression of the recombinant NS1 protein was evaluated by immunofluorescence and flow cytometry. Transfection with either of the two lipid-based methods led to very low number of HepG2 cells expressing NS1 (3.9% and 6.8% with Lipofectamine and FuGene, respectively) and high cell death rates. On the other hand, the efficiency of cell transfection was remarkable higher with nucleofection when compared to these other methods, achieving 63% of cells expressing NS1 protein and more than 60% of viability in the optimized condition.

Keywords


HepG2, Transfection, Nucleofection, Lipid-based methods, NS1, dengue

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DOI: http://dx.doi.org/10.17525/vrrjournal.v23i1.339

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