Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 10^7 HeLa cells and 4 µg of INSERT AB ID [INSERT CLONE ID]. 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 ab214808(red), and 20µl of Anti rabbit IgG sepharose beads. 2μg of rabbit normal IgG was added to the beads control (grey). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.

Immunofluorescent analysis of 100% methanol-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells, untreated and treated with Trichostatin A (500 ng/ml) for 4 hours, labeling Histone H3 (acetyl K64) with ab214808 at 1/2000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cells. 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 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

All lanes : Anti-Histone H3 (acetyl K64) antibody [EPR20713] - ChIP Grade (ab214808) at 1/5000 dilution

Lane 1 : Untreated NIH/3T3 (mouse embyro fibroblast cell line) whole cell lysate
Lane 2 : NIH/3T3 treated with 500ng/ml Trichostatin A for 4 hours whole cell lysate
Lane 3 : Untreated C6 (rat glial tumor cell line) whole cell lysate
Lane 4 : C6 treated with 500ng/ml Trichostatin A for 4 hours whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

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


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

All lanes : Anti-Histone H3 (acetyl K64) antibody [EPR20713] - ChIP Grade (ab214808) at 1/1000 dilution

Lane 1 : Untreated HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : HeLa treated with 400ng/ml Trichostatin A for 9 hours whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

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


Exposure time: 15 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

ab214808 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.