Anti-Histone H2B (mono methyl K85) antibody [EPR17613] (ab177781) at 1/500 dilution + Mouse ovary lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Developed using the ECL technique.

Predicted band size: 14 kDa
Observed band size: 14 kDa


Exposure time: 5 seconds

Blocking/Dilution buffer: 5% BSA/TBST.

Anti-Histone H2B (mono methyl K85) antibody [EPR17613] (ab177781) at 1/5000 dilution + HL-60 (human promyelocytic leukemia cell line) cell lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Developed using the ECL technique.

Predicted band size: 14 kDa
Observed band size: 14 kDa


Exposure time: 5 seconds

Blocking/Dilution buffer: 5% BSA/TBST.

All lanes : Anti-Histone H2B (mono methyl K85) antibody [EPR17613] (ab177781) at 1/1000 dilution

Lane 1 : Human ovary lysate
Lane 2 : HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate
Lane 3 : NIH:OVCAR-3 (human ovary adenocarcinoma cell line) cell lysate
Lane 4 : MOLT-4 (human lymphoblastic leukemia cell line) cell lysate
Lane 5 : THP-1 (human monocytic leukemia cell line) cell lysate
Lane 6 : K562 (human chronic myelogenous leukemia cell line from bone marrow) cell lysate
Lane 7 : F9 (mouse embryonic testicular cancer cell line) cell lysate
Lane 8 : RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) cell lysate
Lane 9 : NIH/3T3 (mouse embyro fibroblast cell line) cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Developed using the ECL technique.

Predicted band size: 14 kDa
Observed band size: 14 kDa


Exposure time: 15 seconds

Blocking/Dilution buffer: 5% BSA/TBST.

Under our testing conditions (using whole cell extract and BSA blocking) we detected non-specific bands in addition to the BOI.

ab177781 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.

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Histone H2B (mono methyl K85) with ab177781 at 1/2000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on HeLa cell line.

The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/500 dilution (red).

The negative controls are as follows:

-ve control 1: ab177781 at 1/2000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/500 dilution.

-ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/500 dilution.