Anti-Histone H3 (phospho S10) antibody [EPR24060-36]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labelling Histone H3 (phospho S10) with ab267372 at 1/1000 (0.54 μg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear staining in HeLa cells in M phase. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunohistochemical analysis of paraffin-embedded human testis tissue labelling Histone H3 (phospho S10) with ab267372 at 1/100 (5.4 μg/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human testis without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab267372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• Flow Cyt

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Flow Cytometry - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (Human cervix adenocarcinoma cell) cells labelling Histone H3 (phospho S10) with ab267372 at 1/500 dilution (0.1μg)/ Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Cells were co-stained with DRAQ5 to differentiate cell cycle phase.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue labelling Histone H3 (phospho S10) with ab267372 at 1/100 (5.4 μg/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human colon carcinoma. The section was incubated with ab267372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IP

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Immunoprecipitation - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Histone H3 (phospho S10) was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell ) treated with 100nM calyculin A for 30 minutes, whole cell lysate 20 μg with ab267372 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab267372 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HeLa treated with 100nM calyculin A for 30 minutes, whole cell lysate 20 μg

Lane 2 : ab267372 IP in HeLa treated with 100nM calyculin A for 30 minutes whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab267372 in HeLa treated with 100nM calyculin A for 30 minutes whole cell lysate

Blocking and dilution buffer and concentration : 5% BSA/TBST

Exposure time : 5.5 seconds

All lanes:

Immunoprecipitation - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (ab267372)

Predicted band size: 15 kDa

false

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 cells labelling Histone H3 (phospho S10) with ab267372 at 1/1000 (0.54 μg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing nuclear staining in NIH/3T3 cells in M phase. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The nuclear counterstain was DAPI (Blue).

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunohistochemical analysis of paraffin-embedded rat testis tissue labelling Histone H3 (phospho S10) with ab267372 at 1/5000 (0.108 μg/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on rat testis without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab267372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Immunohistochemical analysis of paraffin-embedded mouse testis tissue labelling Histone H3 (phospho S10) with ab267372 at 1/5000 (0.108 μg/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on mouse testis without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab267372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• WB

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Western blot - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 9859993).

Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (ab267372) at 1/1000 dilution was used as a control for nocodazole induced cell cycle arrest.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Survivin antibody [RM1086] (<a href='/en-us/products/primary-antibodies/survivin-antibody-rm1086-ab316022'>ab316022</a>) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HeLa treated with 400ng/ml nocodazole for 20 hours 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 1/100000 dilution

Observed band size: 16 kDa,36 kDa,15 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Blocking and diluting buffer and concentration : 5% BSA/TBST

Exposure time : 3 seconds

All lanes:

Western blot - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (ab267372) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell ), whole cell lysate at 20 µg

Lane 2:

HeLa treated with 100nM calyculin A for 30 min, whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast), whole cell lysate at 20 µg

Lane 4:

NIH/3T3 treated with 100nM calyculin A for 30 min, whole cell lysate at 20 µg

Lane 5:

C6 (rat glial tumor glial cell), whole cell lysate at 20 µg

Lane 6:

C6 treated with 100nM calyculin A for 30 min, 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 1/100000 dilution

Predicted band size: 15 kDa

false

• PepArr

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Peptide Array - Anti-Histone H3 (phospho S10) antibody [EPR24060-36] (AB267372)

Peptide array analysis of ab267372 at (0.02μg/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Fluo 647nm conjugated at 1/50,000 dilution.

All batches of ab267372 are 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 the area under the curve when antibody binding values are plotted against the corresponding peptide concentration. Each circle area is normalized to the peptide with the strongest affinity.