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Synthetic peptide conjugated to KLH derived from within residues 100 to the C-terminus of Human p21.
(Peptide available as ab18623.)
Our Abpromise guarantee covers the use of ab18209 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use a concentration of 5 µg/ml.|
|Sandwich ELISA||Use a concentration of 0.5 µg/ml. Can be paired for Sandwich ELISA with Mouse monoclonal [AC8.3] to p21 (ab118). For sandwich ELISA, use this antibody as Detection at 0.5µg/ml with Ab118 as Capture.|
|WB||Use a concentration of 1 µg/ml. Detects a band of approximately 21 kDa (predicted molecular weight: 21 kDa).Can be blocked with Human p21 peptide (ab18623).|
|IHC||Use at an assay dependent concentration. ab171870-Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.|
|IHC-P||Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
IHC image of p21 staining in human lung FFPE section, performed on a 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 ab18209, 5ug/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.
ab18209 staining p21 in serum starved PC-3 cells treated with β-lapachone (ab141097), by ICC/IF. Increase of p21 nuclear expression correlates with increased concentration of β-lapachone, as described in literature.
The cells were incubated at 37°C for 6 hours in media containing different concentrations of ab141097 (β-lapachone) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab18209 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A anti-rabbit DyLight® 488 (ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.
ICC/IF image of ab18209 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab18209, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab18209 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406
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