Use of Confocal Microscopy to Characterise DENV Infection and NS1 Expression in C6/36 and HepG2 Cells in Utilising Them as Models to Screen for the Efficacy of Antiviral Drugs

摘要

Finding receptors, ligands and other molecules which facilitate or inhibit the entry of Dengue virus (DENV) into cells is important, so that they can be manipulated to interrupt viral entry into the cells, hence preventing viral infection. Simultaneously, it is important to identify drug candidates against DENV infection. Confocal laser scanning microscopy can be used to observe viral infection inside cells. Detecting viral infection within cells in the presence and absence of the tested molecules using confocal laser scanning microscopy would be highly important to draw better conclusions. This study is aimed at characterizing DENV infection in C6/36 and HepG2 cell lines, to use them as models to screen the efficacy of antiviral drugs, and to identify detailed disease mechanisms. C6/36 and HepG2 cells were infected with different multiplicity of infections (MOIs: 0.2, 0.4, 1 and 2), and immunofluorescence staining protocols were optimized to choose the optimum antibody concentrations and incubation periods. DENV MOI 0.2 was selected as the optimum concentration to work with, for both C6/36 and HepG2 cell lines. Although 1 hour incubation was sufficient for primary and secondary antibody incubations in immunofluorescence staining for envelope antigen, NS1 staining required an overnight incubation of primary antibody. These protocols may be helpful in identifying inducers and inhibitors of viral entry to C6/36 and HepG2, as well as to experiment on possible indigenous antiviral medicine against dengue.