Transduction efficiency and cytotoxicity of SFV and SIN alphaviral vectors in 4 T1 cells. (a) 4 T1 cells were infected with SFV and SIN particles encoding EGFP. At 24 h post-infection, the cells were harvested, stained with PI and subjected to dual FACS analysis. The x-axis and the y-axis represent EGFP and PI fluorescence, respectively. The percentage of living/dead cells and EGFP-positive/negative cells is indicated on the plot. The FACS data shown are from representative experiments (n = 3). The diagram on the left (MTT assay) demonstrates the cytotoxic effects of SFV and SIN infection. An MTT cell viability assay was performed every day for 5 days post-infection. The results are presented as the percentage of viable cells relative to the control (uninfected cells). The error bars indicate the standard error of 3 independent experiments. (b) Repeated infection of 4 T1 cells. The cells were infected with SFV expressing EGFP (pictures show green fluorescence) and then re-infected 24, 48 and 72 h later with SFV expressing DS-Red (pictures show red fluorescence). Fluorometry of DS-Red fluorescence was performed 1 day after each re-infection. The diagrams represent the percentage of fluorescence units in re-infected cells relative to control cells (100%), which were primarily infected with only SFV/DS-Red. The error bars indicate the standard error of three experiments.