Validation of a HTS assay for IAV transcription/replication inhibitors.

<p>(A) Determination of R² value. A549-5Ps cells were treated with various concentration of ribavirin for 72 hours, followed by Gluc activity analysis. R² was calculated in two independent experiments (each experiment was triplicates). (B) Determination of parameters for the screening assay. A549-5Ps and A549-Gluc cells were seeded into 96-well plates (1×10⁴/well) and cultured for 72h hours. Supernatants were harvested to measure Gluc activity. Z’ factor, signal-to-noise ratio (S/N) and signal-to-background ratio (S/B) were calculated using the following equations: Z’ = 1−3×(SD of A549-5Ps + SD of A549-Gluc)/(mean of A549-5Ps − mean of A549-Gluc); S/N = (mean of A549-5Ps − mean of A549-Gluc)/SD of A549-Gluc; S/B = mean of A549-5Ps/mean of A549-Gluc.