The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml. Detects a band of approximately 77 kDa (predicted molecular weight: 75 kDa).
Use a concentration of 5 µg/ml.
RelevanceThe p53 tumor suppressor gene integrates numerous signals that control cell life and death. There are several proteins that are involved in the p53 pathway, including Chk2, p53R2, p53AIP1, Noxa, PIDD, and MTA2. The transcriptional activity of p53 is modulated by protein stability and acetylation. MTA2, also termed MTA1L1, was found to be a subunit of nucleosome remodeling and deacetylating (NuRD) complex. MTA2 modulates the enzymatic activity of the histone deacetylase complex and its expression reduces the levels of acetylated p53. Deacetylation of p53 by MTA2 represses p53 dependent transcriptional activation and modulates p53 mediated cell growth arrest and apoptosis. MTA2 is ubiquitously expressed in human tissues.
ICC/IF image of ab74489 stained MCF7 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab85872, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) HeLa and Hek293 cells at 5µg/ml.
IHC image of MTA2 staining in human normal cervix formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab85872, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
Western blot - Anti-MTA2 antibody (ab85872)
Anti-MTA2 antibody (ab85872) at 1 µg/ml + PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 10 µg
Secondary Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution developed using the ECL technique