Figure 9.
Evaluation of suitability of the newly developed Cx43 GJ assay for high-throughput screening (HTS). The quality of the assay was assessed by calculating the screening window coefficient (Z´-Factor) as previously described [31]. A coculture of donor cells (90,000 cells/well) and biosensor cells (30,000 cells/well) at a ratio of 3:1 was incubated in assay buffer for 45 min. Prior to the addition of the A2AAR agonist CGS-21680 (1 µM) to the coculture, basal luminescence was measured (red data points, negative control). The coculture was then activated by the addition of CGS-21680 (1 µM). The luminescence signal had reached a stable plateau 19 min after stimulation, and data points were measured as positive controls (green data points). We measured 15 separate data points for positive and negative controls, indicating reproducibility and an assay window of about 3-fold.