Figure 2

Application of CAPPIA for the detection of hormone-dependent interactions. Triplicate slides each carrying 208 features, including all possible combinations of the 16 prey and 10 bait proteins listed in Additional file 1 and sets of positive (p53+SV40T) and negative (p53+TRAF, single baits and preys) controls were reverse transfected with Hek293T cells in the presence of 10 nM R1881 for 3 days. Each prey-bait combination was printed together with the autofluorescent reporter as single spots per slide with a spot to spot distance of 1.5 mm. Following transfection the fluorescence signals of all 624 features were collected and processed as described in methods. The normalised fluorescence signals obtained from the 480 different bait-prey features (10 baits × 16 preys × 3 replicate slides) are shown separately for the different slides. Sample numbers correspond to the combination of bait and prey as summarized in Additional file 1. The cutoff value (indicated as red line) resembles the level of reporter expression obtained with the non-interacting control proteins p53+TRAF. This cross-screening using CAPPIA immediately identifies so-called bait-or prey-unspecific false positives as is exemplified by bait B504 (marked with a star). A specific interaction was identified between B487 and P506 corresponding to the AR-LBD and AR-NTD (sample number 23, marked with an arrow).