Anti-mH2A1 antibody

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mH2A1 antibody (AB37264)

IHC image of mH2A1 staining in human skin FFPE section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab37264, 5μg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-mH2A1 antibody (AB37264)

ab37264 staining mH2A1 in HeLa cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab37264 at 1µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue). Also suitable in cells fixed with 100% methanol (5 min). Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• WB

Lab

Western blot - Anti-mH2A1 antibody (AB37264)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : MH2A1 knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HeLa whole cell lysate (20 μg)
Lane 4 : HepG2 whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab37264 observed at 40 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab37264 was shown to specifically recognize MH2A1 in wild-type HAP1 cells along with additional cross reactive bands. No band was observed when MH2A1 knockout cells were examined. Wild-type and MH2A1 knockout samples were subjected to SDS-PAGE. ab37264 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1 ug/ml and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-mH2A1 antibody (ab37264)

Predicted band size: 40 kDa

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• WB

Unknown

Western blot - Anti-mH2A1 antibody (AB37264)

All lanes:

Western blot - Anti-mH2A1 antibody (ab37264) at 1 µg

Lane 1:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 20 µg

Lane 2:

HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate at 20 µg

Lane 3:

NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/10000 dilution

Predicted band size: 40 kDa

Observed band size: 100 kDa,40 kDa

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Exposure time: 16min