Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• ChIP-seq

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ChIP-sequencing - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• ChIP

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ChIP - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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.

• WB

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Western blot - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

All lanes:

Western blot - 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 at 10 µg

Lane 2:

Untreated HeLa whole cell lysates at 10 µg

Lane 3:

NIH/3T3 (Mouse embryo fibroblast cell line) treated with Trichostatin A 500 ng/ml for 4hr whole cell lysates at 10 µg

Lane 4:

Untreated NIH/3T3 whole cell lysates at 10 µg

Predicted band size: 15 kDa

Observed band size: 15 kDa

false

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (Human epithelial cell line from cervix adenocarcinoma) cells and 5 µg of ab177177 [EPR16988]. 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 H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (Human epithelial cell line from cervix adenocarcinoma) cells and 5 µg of ab177177 [EPR16988]. 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 H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 HeLa (Human epithelial cell line from cervix adenocarcinoma) cells and 5 µg of ab177177 [EPR16988]. 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.

• CUT&Tag-seq

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CUT&Tag sequencing - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

CUT&Tag-seq was performed using 200,000 Oli-neu (Oligodendrocyte progenitor) cells. Cells were permeabilized with 0.05% Digitonin and 0.01% NP-40 for 3 minutes. A 1 : 100 dilution of Recombinant Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (ab177177) was used, along with a Guinea pig anti-rabbit Secondary. DNA was seg using Illumina NovaSeq S Prime to a depth of 22 million reads.

This image is courtesy of Dr Marek Bartosovic, Gonçalo Castelo-Branco Group, Karolinska Institutet.

This experiment and image is courtesy of Dr Marek Bartosovic, Gonçalo Castelo-Branco Group, Karolinska Institutet.

• PepArr

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Peptide Array - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

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, is available under the Product Protocol section.

• CUT&Tag

Supplier Data

CUT&Tag - Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (AB177177)

CUT&Tag-seq was performed using 200,000 Oli-neu (Oligodendrocyte progenitor) cells. Cells were permeabilized with 0.05% Digitonin and 0.01% NP-40 for 3 minutes. A 1 : 100 dilution of Recombinant Anti-Histone H3 (acetyl K9) antibody [EPR16988] - ChIP Grade (ab177177) was used, along with a Guinea pig anti-rabbit Secondary. DNA was seg using Illumina NovaSeq S Prime to a depth of 22 million reads. This image is courtesy of Dr Marek Bartosovic, Gonçalo Castelo-Branco Group, Karolinska Institutet. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

This experiment and image is courtesy of Dr Marek Bartosovic, Gonçalo Castelo-Branco Group, Karolinska Institutet.