All lanes : Anti-Histone H2A (ubiquityl K119) antibody [EPR19800] (ab193203) at 1/1000 dilution

Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate
Lane 3 : C6 (rat glial tumor cell line) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

Predicted band size: 22 kDa
Observed band size: 22 kDa

 

Exposure time : Lane 1:3 minutes; Lane 2:15 seconds; Lane 3: 3 minutes.

Blocking/Dilution buffer: 5% BSA/TBST.

For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here(downloadable copy).

All lanes : Anti-Histone H2A (ubiquityl K119) antibody [EPR19800] (ab193203) at 1/1000 dilution

All lanes : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Developed using the ECL technique.

Predicted band size: 22 kDa
Observed band size: 22 kDa


Exposure time: 10 seconds

Blocking/Dilution buffer: 5% BSA/TBST.

The different result is due to the lysates preparation method. The lysate in lane 1 is prepared by 1%SDS Hot Lysate buffer method. The lysate in lane 2 is prepared by RIPA lysis method. This antibody works in 1%SDS Hot Lysates in WB. 

For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).

ab193203 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 100% methanol-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling Histone H2A (ubiquityl K119) with ab193203 at 1/500 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 antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

Immunofluorescent analysis of 100% methanol-fixed NIH/3T3 (mouse embryo fibroblast cell line) cells labeling Histone H2A (ubiquityl K119) with ab193203 at 1/500 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 NIH/3T3 cell line.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.