Cellular and Biophysical Pipeline for the Screening of Peroxisome Proliferator-Activated Receptor Beta/Delta Agonists: Avoiding False Positives

Figure 4

High-throughput screening assay and statistics. (a) HTS results are expressed by the firefly/Renilla ratio of each compound normalized by positive and negative controls, which were set as 100% and 0%, respectively, for each plate. It is possible to observe that the positive control varied among different plates and wells. Despite the fact that the searched extracts presented low PPARß/δ activation, they still presented significant differences in comparison to the negative controls (signal 7 times higher than the negative control standard deviation). Insert: values for each screened plate presented the high reliability of the data. (b) Confirmatory transactivation assay in triplicate using the possible hit candidates from the previously screened Phytobios library with 1 μM GW0742 as the positive control, 1% DMSO as the negative control (vehicle), and 0.01 mg/mL of the tested extracts. We considered confirmed hit candidate extracts that showed firefly/Renilla ratios seven times higher than the standard deviation (>7 × DP) for the negative control treatment for at least two of the three replicates. Data are the mean ± SD. values were calculated by the unpaired -test (, , and ) with GraphPad Prism.

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