Fig. 1: Comparative characteristics of reporter orthoflaviviruses.
From: Multiplexed multicolor antiviral assay amenable for high-throughput research
a Design of DENV-2 reporter viruses23. Individual reporter viruses are identical except for their respective fluorescent proteins (FPs) inserted as reporters. b HCI readout of cells infected by DENV-2 reporter viruses. Vero cells expressing near-infrared fluorescent protein (V-NIR) infected with respective DENV-2/X-FPs at an MOI of 0.1. Images taken on live cells at 5 days post-infection (dpi). Scale bar: 255 μm. c Design of DENV-2/mAzurite, JEV/eGFP and YF17D/mCherry. Reporter genes flanked by a repeat for the first 21-35 amino acids of the respective capsid protein (C2-35* for DENV-2, C2-34* for JEV, and C2-21* for YF17D) and a ribosome-skipping T2A peptide. d V-NIR infected with DENV-2/mAzurite, JEV/eGFP or YF17D/mCherry. Representative fluorescent images taken by HCI at 3 dpi with an MOI of 0.2. Scale bar: 255 μm. e Kinetics of individual reporter flavivirus infections. V-NIR individually infected by either DENV-2/mAzurite, JEV/eGFP or YF17D/mCherry at an MOI of 0.05, 0.1, 0.2 or 0.5. Fluorescent signals detected daily until 7 dpi by HCI. Cell viability defined as number of NIR positive cells in virus infected wells versus number in the uninfected wells; virus infectivity defined as number of cells with respective fluorescent signals versus number of NIR positive cells in the same well. f Plaque phenotype of DENV-2/mAzurite, JEV/eGFP and YF17D/mCherry. Representative photographs taken from titration of working virus stocks at a dilution of 1:106, 1:107 and 1:105, respectively (see Supplementary Figs 2 and 3 for comparison with parental viruses). g Dynamics of mixed orthoflaviviruses infections. V-NIR infected with mixtures of DENV-2/mAzurite, JEV/eGFP and YF17D/mCherry at equal ratios (1:1:1); at an MOI of 0.05, 0.1, 0.2 or 0.5. NIR and virus-specific fluorescent signals detected by HCI daily until 7 dpi. Total infection rate defined as total number of cells with Azurite, GFP or Cherry signals versus number of NIR positive cells in the same well (black line). Individual columns show the ratios of DENV-2/mAzurite, JEV/eGFP and YF17D/mCherry among all infected cells in the same well at each timepoint. Data represent means ± SD of n = 3 independent experiments. Source data are provided as a Source Data file.
