All lanes : Anti-Histone H3 (mono methyl R17) antibody [EPR17247] (ab177222) at 1/1000 dilution

Lane 1 : HeLa whole cell lysate
Lane 2 : NIH/3T3 whole cell lysate

Lysates/proteins at 10 µg per lane.

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

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

Blocking buffer/Diluting buffer: 5% NFDM/TBST

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labeling Histone H3 (mono methyl R17) with ab177222 at 1/2000 dilution, followed by Goat anti-rabbit Alexa Fluor® 488 (IgG H&L) (ab150077) secondary antibody at 1/400 dilution (green). Confocal image shows nuclear staining on HeLa cells. 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) at 1/500 dilution (red).
The negative controls are as follows:
-ve control 1: ab177222 at 1/2000 dilution followed by ab150120 (goat ant-mouse AlexaFluor®594 secondary) at 1/500 dilution.
-ve control 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.

ab177222 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 liver tissue labeling Histone H3 (mono methyl R17) with ab177222 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution (ab97051). Nuclear staining on Human liver tissue is observed. Counter stained with Hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution.

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 R17) with ab177222 at 1/2000 dilution, followed by Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution (ab97051). Nuclear staining on mouse neuron cells of cerebral cortex tissue is observed. Counter stained with Hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution .

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

Immunohistochemical analysis of paraffin-embedded rat pancreas tissue labeling Histone H3 (mono methyl R17) with ab177222 at 1/2000 dilution, followed by Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution (ab97051). Nuclear staining on rat glandular epithelium of pancreas tissue is observed. Counter stained with Hematoxylin.

Negative control: PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit HRP (IgG H&L) at 1/500 dilution.

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