Anti-Histone H3 (mono methyl K79) 抗体 [EPR17466] - ChIP Grade (ab177183)

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 ab177183 [EPR17466]. 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 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 ab177183 (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 (Taqman approach for active and inactive loci, Sybr green approach for heterochromatic loci).

Blocking/Dilution buffer: 5% NFDM/TBST.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labeling Histone H3 (mono methyl K79) with ab177183 at 1/2000 dilution, followed by Goat anti-rabbit Alexa Fluor® 488 (IgG) (ab150077) secondary antibody at 1/400 dilution (green). Confocal image showing nuclear staining on Hela cell line. 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) at 1/500 dilution (red).
The negative controls are as follows:
1. ab177183 at 1/2000 dilution followed by ab150120 (goat anti-mouse AlexaFluor®594 secondary) at 1/500 dilution.
2. ab7291 (anti-Tubulin mouse mAb) at 1/500 dilution followed by ab150077 (goat anti-rabbit Alexa Fluor®488; IgG H&L) at 1/400 dilution.

ab177183 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 skin tissue labeling Histone H3 (mono methyl K79) with ab177183 at 1/5000 dilution, followed by ab97051 Goat Anti-Rabbit HRP (IgG H&L ) at 1/500 dilution. Nucleus staining on Human skin tissue is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

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

Immunohistochemical analysis of paraffin-embedded Mouse cerebral cortex tissue labeling Histone H3 (mono methyl K79) with ab177183 at 1/5000 dilution, followed by ab97051 Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution. Nucleus staining on mouse cerebral cortex tissue is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

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 cortex tissue labeling Histone H3 (mono methyl K79) with ab177183 at 1/5000 dilution, followed by ab97051 Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution. Nucleus staining on glandular epithelium of rat colon tissue is observed. Counter stained with Hematoxylin.

Negative control: Using PBS instead of primary ab, secondary ab is prediluted HRP Polymer for Rabbit/Mouse IgG.

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