Pilot HTS Screen.

<p>A. Composite heat map of 99 384-well plates used for the pilot screen using the full complement of 10 multiplexed reporter cell lines. B. Heat map of Plate 39 from the pilot screen showing the organization of controls in the first two columns. Wells designated 'N' correspond to either medium alone or DMSO negative controls. Wells 1–11 correspond, respectively, to PAM₂CSK₄ (TLR2 agonist), Poly(I:C) (TLR3 agonist), LPS (TLR4 agonist), MPLA (TLR4 agonist), Flagellin (TLR5 agonist), C4 (pure TLR7 agonist), IMDQ (Dual TLR7/8 agonist), KHP-3-126 (pure TLR8 agonist), ODN-2006 (TLR9 agonist), C₁₂-<i>i</i>E-DAP (NOD1 agonist), Murabutide (NOD2 agonist). All stimuli were used at a final concentration of 5 μg/mL (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0149848#pone.0149848.s001" target="_blank">S1 Fig</a> for structures). Well 12 corresponds to a 'Master-Mix' which combined all the above-mentioned stimuli to yield an effective concentration of 0.45 μg/mL of the individual ligands. C. Distribution of baseline values and signals (a subset is shown for visual clarity). Means and standard deviations were computed for replicates for each of the stimuli from 99 plates from which Z' factors were calculated. Shown also are Gaussian fits of the histograms.