Anti-Histone H2B (mutated E76K) antibody [EPR27298-41]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized 293T cells transfected with a human H2BC4 WT expression vector containing a myc tag (Left) / 293T cells transfected with a human H2BC4 E76K expression vector containing a myc tag (Right) cells labelling Histone H2B (mutated E76K) with ab313789 at 1/50 dilution (1ug)/ Right (Red) compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labeling Histone H2B (mutated E76K) with ab313789 at 1/2000 (0.27 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Negative control : No staining on human cerebrum.The section was incubated with ab313789 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 was performed with citrate buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Immunohistochemical analysis of paraffin-embedded (A) HEK-293T (human tissue labeling Histone H2B (mutated E76K) with ab313789 at 1/2000 (0.27 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Positive staining on (B) HEK-293T transfected with a human H2BC4 E76K expression vector containing a his tag. No staining on (A) HEK-293T transfected with a human H2BC4 (WT) expression vector containing a his tag and (C) HEK-293T transfected with empty vector containing a his tag. The section was incubated with ab313789 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Heat mediated antigen retrieval was performed with citrate buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized 293T (human embryonic kidney epithelial cell) cells labelling Histone H2B (mutated E76K) with ab313789 at 1/4000 (0.135 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green). Confocal image showing nuclear staining in 293T cells transfected with a human H2BC4 E76K expression vector containing a myc tag, and showing no staining in 293T cells transfected with a human wildtype H2BC4 expression vector containing a myc tag. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). Myc-Tag Mouse mAb (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 0.38 ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labeling Histone H2B (mutated E76K) with ab313789 at 1/2000 (0.27 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Negative control : No staining on mouse cerebrum. The section was incubated with ab313789 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 was performed with citrate buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue labeling Histone H2B (mutated E76K) with ab313789 at 1/2000 (0.27 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Negative control : No staining on rat cerebrum. The section was incubated with ab313789 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 was performed with citrate buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

• WB

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Western blot - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Blocking and diluting buffer and concentration : 5% NFDM/TBST The identity of the higher MW band at approximately 37 kDa is unknown. In Western blot, anti-His antibody (ab213204) staining at 1/5000 dilution. In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. Exposure time : 180 seconds

All lanes:

Western blot - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (ab313789) at 1/1000 dilution

Lane 1:

293T cells transfected with an empty vector containing a His-tag whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human H2BC4 E76K expression vector containing a His-tag whole cell lysate at 16 µg

Lane 3:

293T cells transfected with a human H2BC4 (WT) expression vector containing a His-tag whole cell lysate at 20 µg

Secondary

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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: 20 kDa

false

Exposure time: 180s

• Dot

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Dot Blot - Anti-Histone H2B (mutated E76K) antibody [EPR27298-41] (AB313789)

Dot blot analysis of Histone H2B (mutated E76K) using ab313789 at 1 : 1000 (0.512 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution. Lane 1 : Histone H2BE76K peptide Lane 2 : Histone H2BE76Q peptide Lane 3 : Histone H2BG53D peptide Lane 4 : Histone H2BE113K peptide Lane 5 : Histone H2BE76 peptide Exposure time : 180 seconds Blocking and diluting buffer and concentration : 5% NFDM/TBST