Fig 3.

Assay validation for high-throughput inhibitor screening. (A) Control inhibitors with distinct antimyxovirus profiles were reproducibly identified in single-well coinfection assays. IAV-firefly luciferase-transfected cells were infected with IAV-Texas and recMeV-ren in the presence of JMN3-003 (pan-myxovirus inhibitor), AS-136A (MeV RdRp inhibitor), 5-iodotubercidin (IAV inhibitor), or vehicle control (DMSO). Relative luciferase activities in the wells were determined at 30 hpi. Each concentration was assessed in three replicates; two independent plates were prepared and analyzed. Symbols show means (lines) and minimum and maximum values (floating bars). (B) Plate-to-plate variation assessment using a random test set of 480 compounds. Cells were treated at a final concentration of 5 μM and then coinfected, and luciferase activities were determined at 30 hpi. Symbols represent values for each compound obtained from two independent replicate sets. A direct linear correlation (black line), median RLU values for each reporter data set (solid blue and red lines), and 25th and 75th percentiles (dashed blue and red lines) are shown.