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Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] is a rabbit recombinant monoclonal antibody that is used to detect gamma H2A.X in Dot, ICC/IF, IHC-P, IP, Western blot. Suitable for Human, Mouse, Rat samples.

- Using biophysical QC, antibody identity is confirmed at a molecular level for unrivaled batch-batch consistency
- Cited in over 270 publications
- Trusted since 2009


Images

Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (AB81299), expandable thumbnail
  • Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (AB81299), expandable thumbnail
  • Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (AB81299), expandable thumbnail
  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (AB81299), expandable thumbnail
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (AB81299), expandable thumbnail

Publications

Key facts

Isotype
IgG
Host species
Rabbit
Storage buffer

pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA

Form
Liquid
Clonality
Monoclonal

Immunogen

  • The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application
Product promiseTestedExpectedPredictedNot recommended
IPDotWBIHC-PICC/IF
Human
Tested
Expected
Tested
Tested
Tested
Mouse
Expected
Expected
Tested
Not recommended
Expected
Rat
Expected
Expected
Tested
Not recommended
Expected
Sheep
Predicted
Predicted
Predicted
Not recommended
Predicted
Synthetic peptide - Human
Not recommended
Tested
Not recommended
Not recommended
Not recommended

Tested
Tested

Species
Human
Dilution info
1/40
Notes

-

Expected
Expected

Species
Mouse, Rat
Dilution info
Use at an assay dependent concentration.
Notes

-

Predicted
Predicted

Species
Sheep
Dilution info
-
Notes

-

Not recommended
Not recommended

Species
Synthetic peptide - Human
Dilution info
-
Notes

-

Tested
Tested

Species
Synthetic peptide - Human
Dilution info
1/1000
Notes

-

Expected
Expected

Species
Human, Mouse, Rat
Dilution info
Use at an assay dependent concentration.
Notes

-

Predicted
Predicted

Species
Sheep
Dilution info
-
Notes

-

Tested
Tested

Species
Human
Dilution info
1/1000 - 1/500000
Notes

-

Species
Mouse
Dilution info
1/1000 - 1/500000
Notes

-

Species
Rat
Dilution info
1/1000 - 1/500000
Notes

-

Predicted
Predicted

Species
Sheep
Dilution info
-
Notes

-

Not recommended
Not recommended

Species
Synthetic peptide - Human
Dilution info
-
Notes

-

Tested
Tested

Species
Human
Dilution info
-
Notes

-

Not recommended
Not recommended

Species
Mouse, Rat, Sheep, Synthetic peptide - Human
Dilution info
-
Notes

-

Tested
Tested

Species
Human
Dilution info
1/250
Notes

-

Expected
Expected

Species
Mouse, Rat
Dilution info
Use at an assay dependent concentration.
Notes

-

Predicted
Predicted

Species
Sheep
Dilution info
-
Notes

-

Not recommended
Not recommended

Species
Synthetic peptide - Human
Dilution info
-
Notes

-

Associated Products

Select an associated product type

6 products for Alternative Product

Target data

Function

The protein expressed by the H2AX gene is a variant histone H2A that replaces conventional H2A in certain nucleosomes, which are responsible for wrapping and compacting DNA into chromatin. This compaction limits DNA accessibility to cellular machineries that require DNA as a template, placing histones at the center of transcription regulation, DNA repair, DNA replication, and chromosomal stability. DNA accessibility is controlled through a complex array of post-translational histone modifications, known as the histone code, and nucleosome remodeling. The H2AX protein is essential for the checkpoint-mediated arrest of cell cycle progression in response to low doses of ionizing radiation and for the efficient repair of DNA double strand breaks (DSBs), particularly when it undergoes C-terminal phosphorylation. This supplementary information is collated from multiple sources and compiled automatically.

Alternative names

H2AFX, H2AX, Histone H2AX, H2a/x, Histone H2A.X, H2AS139p, H2AXS139p, H2A.XS139p

Recommended products

Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] is a rabbit recombinant monoclonal antibody that is used to detect gamma H2A.X in Dot, ICC/IF, IHC-P, IP, Western blot. Suitable for Human, Mouse, Rat samples.

- Using biophysical QC, antibody identity is confirmed at a molecular level for unrivaled batch-batch consistency
- Cited in over 270 publications
- Trusted since 2009

Key facts

Isotype
IgG
Form
Liquid
Clonality
Monoclonal
Immunogen
  • The exact immunogen used to generate this antibody is proprietary information.
Clone number
EP854(2)Y
Purification technique
Affinity purification Protein A
Concentration
Loading...

Storage

Shipped at conditions
Blue Ice
Appropriate short-term storage duration
1-2 weeks
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
-20°C
Aliquoting information
Upon delivery aliquot
Storage information
Avoid freeze / thaw cycle

Notes

Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) was developed by Abcam using patented rabbit monoclonal antibody technology and is validated for use in Dot Blot, ICC/IF, IHC-P, IP and WB.

Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) was first used in a scientific publication in 2008 and has been cited over 272 times in peer reviewed journals. It's performance in Western Blot and immunofluorescence in human samples is trusted by the scientific community.

Abcam's high quality manufacturing and validation processes ensure Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) has high sensitivity and specificity alongside high lot-to-lot consistency and reproducibility.

Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) has 16 independent reviews from customers.

Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) specifically detects gamma H2A.X Phospho-S139 (UniProt ID: P16104; Molecular weight: 16kDa) and is sold in a convenient trial size to enable initial testing (20 µL) and larger sizes for subsequent scaling up experiments (100 µL and 1 mL).

Conjugation-ready, carrier free format available for antibody clone EP854(2)Y - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] - BSA and Azide free ab215967.

Antibody clone EP854(2)Y is also available pre-conjugated to a variety of labels for your convenience - Alexa Fluor® 488, Alexa Fluor® 647, HRP, Alexa Fluor® 594, Alexa Fluor® 555 (Alexa Fluor® 488 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195188, Alexa Fluor® 647 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195189, HRP Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab195190, Alexa Fluor® 594 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab206898, Alexa Fluor® 555 Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] ab206900).

Gamma H2A.X (γH2A.X) is a phosphorylated form of the H2A histone family member X (H2AX), which plays a crucial role in the cellular response to DNA damage. When double-strand breaks occur in DNA, H2AX is rapidly phosphorylated at serine 139 to form γH2A.X. This modification serves as a marker for DNA damage and is essential for recruiting DNA repair proteins to the site of damage. Researchers use γH2A.X as a sensitive indicator of DNA double-strand breaks in studies of genotoxic stress, cancer and other conditions involving DNA damage and repair.

Patented technology
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

What are the advantages of a recombinant monoclonal antibody?
This product is a recombinant monoclonal antibody, which offers several advantages including:

  • - High batch-to-batch consistency and reproducibility
  • - Improved sensitivity and specificity
  • - Long-term security of supply
  • - Animal-free batch production

For more information, read more on recombinant antibodies.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.
Activity summary

Gamma H2A.X also known as phospho H2A.X or γH2A.X is a phosphorylated form of the histone variant H2A.X. It has a molecular weight of about 14 kilodaltons and occurs primarily in they nucleus. When DNA double-strand breaks (DSBs) occur serine 139 in H2A.X undergoes rapid phosphorylation resulting in gamma H2A.X. This modification happens swiftly at the site of damage and gamma H2A.X spreads over a large chromatin area facilitating the recruitment of DNA repair proteins. Gamma H2A.X staining typically evaluated using gamma H2A.X immunofluorescence techniques aids in identifying the presence and extent of DNA damage.

Biological function summary

Gamma H2A.X plays a role in DNA damage response and repair. It does not operate alone; it acts as part of a complex with other repair proteins. The formation of gamma H2A.X foci at DNA damage sites creates a signal attracting repair factors that help maintain genome stability. Its interaction with MDC1 and ATM proteins exemplifies its significant role in orchestrating an effective response to DNA damage. Beyond DNA repair gamma H2A.X influences cell cycle checkpoints permitting cells to pause and repair before proceeding with division.

Pathways

Gamma H2A.X plays a pivotal role in the DNA damage response (DDR) pathway. This pathway is essential for detecting and repairing DNA lesions to uphold genomic integrity. Within the DDR pathway gamma H2A.X is closely associated with proteins such as NBS1 and BRCA1 which assist in repairing double-strand breaks. In addition gamma H2A.X is integral to the ATM-ATR signaling pathway where its activation promotes cell survival following genotoxic stress by signaling for damage repair or triggering apoptosis.

Associated diseases and disorders

Gamma H2A.X has connections to cancer and neurodegeneration. Aberrant DNA repair pathways often indicated by persistent gamma H2A.X signals correlate with tumor formation and progression. For instance a failure to repair DNA damage effectively can lead to mutations that drive cancer development. Gamma H2A.X also links to neurodegenerative diseases where dysregulated DNA repair contributes to neuronal cell death. Proteins like p53 which regulate cell cycle and apoptosis further connect to gamma H2A.X bridging its role in disease pathogenesis.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

17 product images

  • Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunocytochemistry/Immunofluorescence analysis of HeLa cells (untreated and treated with H2O2) labelling Histone H2A.X (phospho S139) with ab81299 at 1/250. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. The cells were co-stained with Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291, a mouse anti-tubulin (1/1000) using Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) as the secondary antibody. Nuclei counterstained with DAPI (blue).

    Control 1: primary antibody (1/250) and secondary antibody, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).

    Control 2: Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291 (1/1000) and secondary antibody, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).

  • Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunocytochemistry/ Immunofluorescence - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunofluorescence staining of A549 (Human lung carcinoma cell line) labeling gamma H2A.X (phospho S139) (green) with ab81299.

    Cells were fixed in 4% paraformaldehyde for 15 minutes and permeabilized in PBS-0.2% Triton for 10 minutes. After blocked for 1 hour, primary antibody was diluted in blocking buffer (1/100) and incubated with fixed cells overnight at 4°C. Cells were washed and incubated with secondary antibodies (1/100) for 1 hour at room temperature. All slides were counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Immunofluorescence was performed using confocal laser scanning microscopy (Lecia) or fluorescence microscopy (Olympus).

    Dexamethasone, DEX; Cisplatin, DDP.

  • Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    ab81299 at 1/40 immunoprecipitating Histone H2A.X (phospho S139) in HepG2 (human hepatocellular carcinoma epithelial) whole cell lysate observed at 15 KDa (lanes 1 and 2).

    Lane 1 (input): HepG2 treated with etoposide and TSA whole cell lysate 10μg
    Lane 2 (+): ab81299 + HepG2 treated with etoposide and TSA whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab81299 in HepG2 treated with etoposide and TSA

    For western blotting, ab81299 (Purified) at 1/200 dilution and VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366), was used for detection at 1/1000 dilution.

    Blocking buffer and concentration: 5% NFDM/TBST.
    Diluting buffer and concentration: 5% NFDM /TBST.

    All lanes: Immunoprecipitation - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Predicted band size: 15 kDa

    Exposure time: 30s

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Blocking buffer - 5% NFDM/TBST

    Diluting buffer - 1% BSA

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/100000 dilution

    Lane 1: HepG2 cell lysate – treated with etoposide at 20 µg

    Lane 2: HepG2 cell lysate – untreated at 20 µg

    Secondary

    All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

    Predicted band size: 15 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail
    This image is courtesy of a customer review submitted by Carl Hobbs.

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    ab81299 staining H2A.X in Human Testis tissue sections by Immunohistochemistry (Formalin/PFA fixed paraffin embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 10 minutes at 21°C; antigen retrieval was by heat mediation in Citric acid. Samples were incubated with primary antibody (1/50 in TBS) for 2 hours at 21°C. A biotin conjugated Anti-Rabbit IgG (goat polyclonal) was used as the secondary antibody at a 1/250 dilution.

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/500000 dilution

    Lane 1: Jurkat cell lysate - untreated at 10 µg

    Lane 2: Jurkat cell lysate - treated with etoposide at 10 µg

    Secondary

    All lanes: HRP Labelled goat anti-rabbit at 1/2000 dilution

    Predicted band size: 15 kDa

    Observed band size: 15 kDa

  • Dot Blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Dot Blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Dot blot analysis of Histone H2A.X single phospho peptide pS139 (lane 1) and Histone H2A.X non-phospho peptide (lane 2) with ab81299 at 1/1000. Blocking and diluting buffer was 5% NFDM/TBST. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) ab97051 Peroxidase conjugated Goat Anti-Rabbit IgG, (H+L) at 1/100,000.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunohistochemical staining of paraffin embedded human brain with unpurified ab81299 at a working dilution of 1 in 50. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunohistochemical staining of paraffin embedded human brain with purified ab81299 at a working dilution of 1 in 100. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunohistochemical analysis of formalin/PFA-fixed paraffin-embedded human kidney transitional cell carcinoma using unpurified ab81299 at a dilution of 1/100.

    Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue labeling gamma H2A.X with ab81299 at 1/3000 (0.352 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human breast carcinoma without lambda protein phosphatase treatment (image A). No signal was detected when tissues were treated with lambda protein phosphatase (image B). The section was incubated with ab81299 for 15 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 Rabbit specific IHC polymer detection kit HRP/DAB ab209101. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Gamma H2A.X (phospho S139) Western blot staining using rabbit Anti-gamma H2A.X (phospho S139) antibody

    In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining at 1/1000000 dilution.

    In Western blot, Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker (Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker ab124781) staining at 1/2000 dilution.

    The identity of the bands between 20kDa and 75kDa are unknown.

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/1000 dilution

    Lane 1: Untreated PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

    Lane 2: PC-12 treated with 3µM etoposide for 1-hour whole cell lysate at 20 µg

    Lane 3: PC-12 treated with 3µM etoposide for 1-hour whole cell lysate 20µg, then the membrane treated with Alkaline Phosphatase for 1 hour at 20 µg

    Secondary

    All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

    Observed band size: 15 kDa

    Exposure time: 3s

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Gamma H2A.X (phospho S139) Western blot staining using rabbit Anti-gamma H2A.X (phospho S139) antibody

    In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining at 1/1000000 dilution.

    In Western blot, Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker (Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker ab124781) staining at 1/2000 dilution.

    The identity of the bands between 37kDa and 50kDa are unknown.

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/1000 dilution

    Lane 1: Untreated NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 20 µg

    Lane 2: NIH/3T3 treated with 30µg/ml etoposide for 4 hours, 500ng/ml Trichostatin A(TSA) was then added for additional 4 hours whole cell lysate at 20 µg

    Lane 3: NIH/3T3 treated with 30µg/ml etoposide for 4 hours, 500ng/ml Trichostatin A(TSA) was then added for additional 4 hours whole cell lysate 20µg, then the membrane treated with Alkaline Phosphatase for 1 hour at 20 µg

    Secondary

    All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

    Observed band size: 15 kDa

    Exposure time: 180s

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Western blot: Rabbit Monoclonal [EP854(2)Y] to gamma H2A.X (phospho S139) ab81299 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 15 kDa in HepG2 Vehicle Control Etoposide (0 uM, 90 min) cell lysates with no signal observed at this size in HepG2 Etoposide treated (100 uM, 90 min) cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc BSA in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/1000 dilution

    Lane 1: HepG2 Vehicle Control Etoposide (0 uM, 90 min) at 20 µg

    Lane 2: HepG2 Etoposide treated (100 uM, 90 min) at 20 µg

    Lane 3: Jurkat Vehicle Control Etoposide (0 uM, 90 min) at 20 µg

    Lane 4: Jurkat Treated Etoposide (100 uM, 90 min) at 20 µg

    Lane 5: Human Brain at 20 µg

    Secondary

    All lanes: Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

    Performed under reducing conditions.

    Predicted band size: 15 kDa

    Observed band size: 15 kDa

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Mouse anti GAPDH was used as a GAPDH loading control.

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/1000 dilution

    Lane 1: HepG2 Vehicle Control Etoposide (0 uM, 90 min) at 20 µg

    Lane 2: HepG2 Etoposide treated (100 uM, 90 min) at 20 µg

    Lane 3: Jurkat Vehicle Control Etoposide (0 uM, 90 min) at 20 µg

    Lane 4: Jurkat Treated Etoposide (100 uM, 90 min) at 20 µg

    Lane 5: Human Brain at 20 µg

    Predicted band size: 15 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Immunohistochemical analysis of paraffin-embedded Human cervical carcinoma tissue labeling gamma H2A.X with ab81299 at 1/3000 (0.352 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Nuclear staining on human cervical carcinoma without lambda protein phosphatase treatment (image A). No signal was detected when tissues were treated with lambda protein phosphatase (image B). The section was incubated with ab81299 for 15 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 Rabbit specific IHC polymer detection kit HRP/DAB ab209101. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

  • Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299), expandable thumbnail

    Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299)

    Gamma H2A.X (phospho S139) Western blot staining using rabbit Anti-gamma H2A.X (phospho S139) antibody

    In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) staining at 1/1000000 dilution.

    In Western blot, Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker (Anti-Histone H2A.X antibody [EPR895] - Nuclear Marker ab124781) staining at 1/2000 dilution.

    The identity of the bands between 25kDa and 50kDa are unknown.

    All lanes: Western blot - Anti-gamma H2A.X (phospho S139) antibody [EP854(2)Y] (ab81299) at 1/1000 dilution

    Lane 1: Untreated Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate at 20 µg

    Lane 2: Jurkat treated with 25µM etoposide for 8 hours whole cell lysate at 20 µg

    Lane 3: Jurkat treated with 25µM etoposide for 8 hours whole cell lysate 20µg, then the membrane treated with Alkaline Phosphatase for 1 hour at 20 µg

    Secondary

    All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

    Observed band size: 15 kDa

    Exposure time: 60s

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