Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 10⁷ HeLa cells and 4 µg of ab177177 [EPR16988]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads.

Additional screenshots of mapped reads can be downloaded here.

Chromatin was prepared from HeLa (Human epithelial cell line from cervix adenocarcinoma) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab177177 (blue), and 20µl 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 for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

All lanes : Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (ab177177) at 1/1000 dilution

Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) treated with Trichostatin A 500 ng/ml for 4hr whole cell lysates
Lane 2 : Untreated HeLa whole cell lysates
Lane 3 : NIH/3T3 (Mouse embryo fibroblast cell line) treated with Trichostatin A 500 ng/ml for 4hr whole cell lysates
Lane 4 : Untreated NIH/3T3 whole cell lysates

Lysates/proteins at 10 µg per lane.

Predicted band size: 15 kDa
Observed band size: 15 kDa

ab177177 was tested in Peptide Array against 501 different modified and unmodified histone peptides; each peptide is printed on the array at six concentrations (each in triplicate).
Circle area represents affinity between the antibody and a peptide: all antigen-containing peptides are displayed as red circles, all other peptides as blue circles. The affinity is calculated as area under curve when antibody binding values are plotted against the corresponding peptide concentration. Each circle area is normalized to the peptide with the strongest affinity.
The complete dataset, including full list of all peptides and information on the position of each peptide in the diagram, can be downloaded here.

Immunohistochemical analysis of paraffin-embedded human colon tissue labeling Histone H3 (acetyl K9) with ab177177 at 1/500 dilution, followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) at 1/500 dilution. Nuclear staining on glandular epithelium of Human colon tissue is observed. Counterstained with hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded mouse colon tissue labeling Histone H3 (acetyl K9) with ab177177 at 1/500 dilution, followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) at 1/500 dilution. Nuclear staining on glandular epithelium of mouse colon tissue is observed. Counterstained with hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded rat spleen tissue labeling Histone H3 (acetyl K9) with ab177177 at 1/500 dilution, followed by Goat Anti-Rabbit HRP (IgG H&L) (ab97051) at 1/500 dilution. Nuclear staining on rat spleen tissue is observed. Counterstained with hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Histone H3 (acetyl K9) with ab177177 at 1/3000 dilution, followed by Goat anti-rabbit Alexa Fluor^® 488 (IgG) (ab150077) secondary antibody at 1/400 dilution (green). Confocal image shows nuclear staining on HeLa cell line. The expression increased after treatment with Trichostatin A (500 ng/ml) for 4 hours. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution and ab150120 (goat anti-mouse AlexaFluor^®594 secondary antibody) at 1/500 dilution (red).
The negative controls are as follows:
-ve control 1: ab177177 at 1/3000 dilution followed by ab150120 (AlexaFluor^®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution followed by ab150077 (Alexa Fluor^®488 Goat Anti-Rabbit IgG H&L) at 1/400 dilution.