Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells, treated with Trichostatin A (500 ng/ml) for 4 hours or untreated, labeling Histone H2B (acetyl K20) with ab177430 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 is observed. 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;
1. ab177430 at 1/2000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling Histone H2B (acetyl K20) with ab177430 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nucleus staining on Human colon is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody.

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

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Intracellular flow cytometric analysis of 2% paraformaldehyde-fixed HeLa (Human epithelial cells from cervix adenocarcinoma) cells treated with 500ng/ml Trichostatin A for 4 hours, labeling Histone H2B (acetyl K20) with ab177430 at 1/300 dilution (red) compared with a rabbit monoclonal IgG isotype control (black) and a unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/150 dilution was used as the secondary antibody.

• ChIP

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ChIP - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

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 ab177430 (blue), and 20μl of Anti rabbit IgG sepharose beads. 2μg of rabbit normal IgG was added to the beads control (yellow). 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.

• ChIP-seq

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ChIP-sequencing - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

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 ab177430. 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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Immunohistochemical analysis of paraffin-embedded Rat colon tissue labeling Histone H2B (acetyl K20) with ab177430 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nucleus staining on Rat colon is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody.

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Immunohistochemical analysis of paraffin-embedded Mouse kidney tissue labeling Histone H2B (acetyl K20) with ab177430 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Nucleus staining on mouse kidney is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody.

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

• WB

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Western blot - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (ab177430) at 1/10000 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:

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

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated

Predicted band size: 14 kDa

Observed band size: 14 kDa

false

Exposure time: 3min

• WB

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Western blot - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (ab177430) at 1/1000 dilution

Lane 1:

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

Lane 2:

NIH/3T3 (Mouse embyro fibroblast cells) untreated whole cell lysates 10ug

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: 10s

• PepArr

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Peptide Array - Anti-Histone H2B (acetyl K20) antibody [EPR859] - ChIP Grade (AB177430)

ab177430 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.