Synthetic peptide corresponding to Histone H3 aa 1-100 (acetyl K18) conjugated to Keyhole Limpet Haemocyanin (KLH).
(Peptide available as
Our Abpromise guarantee covers the use of ab1191 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/1000. Detects a band of approximately 17 kDa.|
|PepArr||Use a concentration of 0.2 - 0.02 µg/ml.|
|CHIPseq||Use at an assay dependent concentration.|
|ICC/IF||Use a concentration of 1 µg/ml.|
|ChIP/Chip||Use at an assay dependent concentration.|
|ChIP||Use 2µg for 106 cells.|
All batches of ab1191 are tested in Peptide Array against peptides to different Histone H3 and H4 modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H3 - acetyl K18 peptide (ab24003), indicating that this antibody specifically recognises the Histone H3 - acetyl K18 modification.
ab17163 - Histone H3 - unmodified
ab16635 - Histone H3 - acetyl K9
ab15591 - Histone H3 - acetyl K14
ab24003 - Histone H3 - acetyl K18
ab48359 - Histone H3 - acetyl K23
ab24404 - Histone H3 - acetyl K27
ab41409 - Histone H3 - acetyl K36
ab15662 - Histone H4 - acetyl K12
Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 min. The ChIP was performed with 25 µg of chromatin, 2 µg of ab1191 (blue), and 20 µl of protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.
IHC image of Histone H3 (acetyl K18) staining in Human normal skin 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 ab1191, 1µg/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.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.