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 ab177185 [EPR17468]. 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 cells from cervix adenocarcinoma) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The ChIP was performed with 25μg of chromatin, 2μg of ab177185 (blue), and 20μl of Anti rabbit IgGsepharose beads. 2μg of Rabbit normal IgG was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (SYBR approach). Primers and probes are located in the first kb of the transcribed region.

Anti-Histone H3 (mono+di+tri methyl K79) antibody [EPR17468] - ChIP Grade (ab177185) at 1/1000 dilution + HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

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


Exposure time: 30 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

Anti-Histone H3 (mono+di+tri methyl K79) antibody [EPR17468] - ChIP Grade (ab177185) at 1/1000 dilution + NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

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


Exposure time: 1 minute

Blocking/Dilution buffer: 5% NFDM/TBST.

ab177185 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 (mono+di+tri methyl K79) using ab177185 at 1/4000 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Counter stained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab177185.
Note: Nucleus staining on glandular epithelium of Human colon tissue was observed.

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 (mono+di+tri methyl K79) using ab177185 at 1/4000 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Counter stained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab177185.
Note: Nucleus staining on glandular epithelium of Mouse colon tissue was observed.

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

Immunohistochemical analysis of paraffin-embedded Rat colon tissue labeling Histone H3 (mono+di+tri methyl K79) using ab177185 at 1/4000 dilution. A Goat Anti-Rabbit IgG H&L (HRP) (ab97051) was used as secondary at 1/500 dilution. Counter stained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab177185.
Note: Nucleus staining on glandular epithelium of Rat colon tissue was observed.

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