Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Immunofluorescent analysis of 4% paraformaldehyde-fixed 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells untreated and treated with Trichostatin A (500 ng/ml) for 4 hours labeling Histone H2A (acetyl K9) with ab177312 at 1/2000 dilution followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Confocal image showing nuclear staining on HeLa cell line. Acetylation level 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 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows :
-ve control 1 : ab177312 at 1/2000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.

• ChIP-seq

Unknown

ChIP-sequencing - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

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 ab177312 (EPR17471). 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.

• WB

Supplier Data

Western blot - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (ab177312) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cells from cervix adenocarcinoma) treated with 500 ng/ml Trichostatin A for 4 hours whole cell lysates at 10 µg

Lane 2:

Untreated HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysates at 10 µg

Secondary

All lanes:

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

Predicted band size: 14 kDa

Observed band size: 14 kDa

false

Exposure time: 30s

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (human cervical adenocarcinoma epithelial cell) cells and 5 µg of ab177312 [EPR17471]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown.
The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (human cervical adenocarcinoma epithelial cell) cells and 5 µg of ab177312 [EPR17471]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown.
The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (human cervical adenocarcinoma epithelial cell) cells and 5 µg of ab177312 [EPR17471]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown.
The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• WB

Supplier Data

Western blot - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (ab177312) at 1/5000 dilution

Lane 1:

NIH/3T3 (Mouse embyro fibroblast cells) treated with 500 ng/ml Trichostatin A for 4 hours whole cell lysates at 10 µg

Lane 2:

Untreated NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysates at 10 µg

Secondary

All lanes:

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

Predicted band size: 14 kDa

Observed band size: 14 kDa

false

Exposure time: 30s

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Immunohistochemical analysis of paraffin-embedded rat colon tissue labeling Histone H2A (acetyl K9) with ab177312 at 1/4000 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary antibody at 1/500 dilution. Nucleus staining on rat colon tissue is observed. Counter stained with Hematoxylin.

Negative control : Used 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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Immunohistochemical analysis of paraffin-embedded mouse liver tissue labeling Histone H2A (acetyl K9) with ab177312 at 1/4000 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary antibody at 1/500 dilution. Nucleus staining on mouse liver tissue is observed. Counter stained with Hematoxylin.

Negative control : Used 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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling Histone H2A (acetyl K9) with ab177312 at 1/4000 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary antibody at 1/500 dilution. Nucleus staining on Human colon tissue is observed. Counter stained with Hematoxylin.

Negative control : Used 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.

• ChIP

Unknown

ChIP - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

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 10 minutes. The ChIP was performed with 25μg of chromatin 2μg of ab177312 (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 (Sybr green approach). Primers and probes are located in the first kb of the transcribed region.

• PepArr

Supplier Data

Peptide Array - Anti-Histone H2A (acetyl K9) antibody [EPR17471] - ChIP Grade (AB177312)

ab177312 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, is available under the Product Protocol section.