Example positive control sample standard curve. A dilution series of extract in Incubation Buffer in the working range of the assay. The extract was prepared from MCF7 cells treated with camptothecin and trichostatin A (TCA).
Figure 2. The p53 Acetyl K305 ELISA (ab139411) specifically measures the acetylated Lysine305. MCF7 cells were treated with histone deacetylase inhibitor trichostatin A (TCA), sirtuin 1 inhibitor Ex527, NAD+-dependent deacetylase inhibitor nicotinamide (Nico), or drugs’ vehicle (Veh) in the presence of camptothecin (Campto, in red) or drug’s vehicle (DMSO, in green). Acetyl Lysine305 and total p53 protein levels were determined, respectively, using this kit and ab117995. Dilutions of extract of MCF7 cells treated with camptothecin and TCA were used to construct the standard curves.
The detector antibody used in this kit specifically detects the acetylated p53 as determined by Western blotting. MCF7 cells were treated with drugs’ vehicle (DMSO, lane 2), trichostatin A (lane 3), Ex527 (lane 4), nicotinamide (lane 5), camptothecin (lane 6), trichostatin A and camptothecin (lane 7), Ex527 and camptothecin (lane 8), nicotinamide and camptothecin (lane 9). Cell extracts (20 g/ lane) were analyzed by Western blotting using the p53 Acetyl K305 Detector Antibody (A), the p53 capture antibody of this kit to detect total p53 protein (B) and IRDyes labeled secondary antibodies. The overlay of the green signal of p53 Acetyl K305 Detector Antibody with the red signal p53 capture antibody is shown in C.