Hit-antagonist identification during pilot screen and confirmation after retesting.

<p>A, <i>left panel</i>. Realtime monitoring of OT-induced Ca²⁺-mobilization from UT-myo cells. The two end columns (one highlighted in pink) of each plate were used for checkerboard analyses to determine the Z´-factor for each HTS assay. Examples of “Hit”-antagonists are highlighted (blue boxes). A, <i>right panel</i>. OT-induced Ca²⁺-mobilization of highlighted wells. White arrow indicates the time of OT addition. B. Plate heatmap of Ca²⁺-mobilization at the time indicated by the dashed line (right panel A). Representative hit-antagonist compounds are highlighted by blue boxes. C. The average cutoff threshold based on 3*MAD from median (refer to “Materials and Methods” section) for “hit”-antagonists was 41.07±3.79% inhibition and “hit”-agonists (OT-potentiators) was 50.02±4.77% activation. D. Representative heatmap of additional Ca²-mobilization assays performed to retest hit-agonists and antagonists in duplicate. Veh = vehicle, Atos = atosiban