Anti-gamma H2A.X (phospho S139) antibody [9F3] (ab26350)


  • Product name
    Anti-gamma H2A.X (phospho S139) antibody [9F3]
    See all gamma H2A.X primary antibodies
  • Description
    Mouse monoclonal [9F3] to gamma H2A.X (phospho S139)
  • Host species
  • Tested applications
    Suitable for: Flow Cyt, WB, IP, ICC/IF, IHC-P, In situ hybridizationmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Sheep, Rabbit, Chicken, Guinea pig, Hamster, Cow, Dog, Human, Pig, Monkey
  • Immunogen

    Synthetic peptide of phosphorylated (Ser139) human Histone H2A.X.

  • Positive control
    • HeLa heat shocked cell lysate for Western blot.



Our Abpromise guarantee covers the use of ab26350 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Flow Cyt Use 1µg for 106 cells.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 16 kDa.
IP Use a concentration of 12.5 µg/ml.
ICC/IF 1/500.
IHC-P Use a concentration of 1 µg/ml.
In situ hybridization Use at an assay dependent dilution.


  • Function
    Variant histone H2A which replaces conventional H2A in a subset of nucleosomes. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. Required for checkpoint-mediated arrest of cell cycle progression in response to low doses of ionizing radiation and for efficient repair of DNA double strand breaks (DSBs) specifically when modified by C-terminal phosphorylation.
  • Sequence similarities
    Belongs to the histone H2A family.
  • Developmental stage
    Synthesized in G1 as well as in S-phase.
  • Domain
    The [ST]-Q motif constitutes a recognition sequence for kinases from the PI3/PI4-kinase family.
  • Post-translational
    Phosphorylated on Ser-140 (to form gamma-H2AFX or H2AX139ph) in response to DNA double strand breaks (DSBs) generated by exogenous genotoxic agents and by stalled replication forks, and may also occur during meiotic recombination events and immunoglobulin class switching in lymphocytes. Phosphorylation can extend up to several thousand nucleosomes from the actual site of the DSB and may mark the surrounding chromatin for recruitment of proteins required for DNA damage signaling and repair. Widespread phosphorylation may also serve to amplify the damage signal or aid repair of persistent lesions. Phosphorylation of Ser-140 (H2AX139ph) in response to ionizing radiation is mediated by both ATM and PRKDC while defects in DNA replication induce Ser-140 phosphorylation (H2AX139ph) subsequent to activation of ATR and PRKDC. Dephosphorylation of Ser-140 by PP2A is required for DNA DSB repair. In meiosis, Ser-140 phosphorylation (H2AX139ph) may occur at synaptonemal complexes during leptotene as an ATM-dependent response to the formation of programmed DSBs by SPO11. Ser-140 phosphorylation (H2AX139ph) may subsequently occurs at unsynapsed regions of both autosomes and the XY bivalent during zygotene, downstream of ATR and BRCA1 activation. Ser-140 phosphorylation (H2AX139ph) may also be required for transcriptional repression of unsynapsed chromatin and meiotic sex chromosome inactivation (MSCI), whereby the X and Y chromosomes condense in pachytene to form the heterochromatic XY-body. During immunoglobulin class switch recombination in lymphocytes, Ser-140 phosphorylation (H2AX139ph) may occur at sites of DNA-recombination subsequent to activation of the activation-induced cytidine deaminase AICDA. Phosphorylation at Tyr-143 (H2AXY142ph) by BAZ1B/WSTF determines the relative recruitment of either DNA repair or pro-apoptotic factors. Phosphorylation at Tyr-143 (H2AXY142ph) favors the recruitment of APBB1/FE65 and pro-apoptosis factors such as MAPK8/JNK1, triggering apoptosis. In contrast, dephosphorylation of Tyr-143 by EYA proteins (EYA1, EYA2, EYA3 or EYA4) favors the recruitment of MDC1-containing DNA repair complexes to the tail of phosphorylated Ser-140 (H2AX139ph).
    Monoubiquitination of Lys-120 (H2AXK119ub) by RING1 and RNF2/RING2 complex gives a specific tag for epigenetic transcriptional repression. Following DNA double-strand breaks (DSBs), it is ubiquitinated through 'Lys-63' linkage of ubiquitin moieties by the E2 ligase UBE2N and the E3 ligases RNF8 and RNF168, leading to the recruitment of repair proteins to sites of DNA damage. Monoubiquitination and ionizing radiation-induced 'Lys-63'-linked ubiquitination are distinct events.
  • Cellular localization
    Nucleus. Chromosome.
  • Information by UniProt
  • Database links
  • Alternative names
    • H2A histone family member X antibody
    • H2A histone family member X antibody
    • H2A.FX antibody
    • H2A.X antibody
    • H2a/x antibody
    • H2AFX antibody
    • H2AX antibody
    • H2AX_HUMAN antibody
    • Histone H2A.X antibody
    see all


  • ab26350 (1/500) staining gamma H2A.X (phospho S139) in HeLa cells (green). Cells were fixed in paraformaldehyde, permeabilised with 0.5% Triton X-100/PBS and counterstained with DAPI in order to highlight the nucleus (red). For further experimental details please refer to Abreview.

    See Abreview

  • ab26350 (1µg/ml) staining gamma H2A in human spleen, using an automated system (DAKO Autostainer Plus). Using this protocol there is strong nuclear staining .
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
  • All lanes : Anti-gamma H2A.X (phospho S139) antibody [9F3] (ab26350)

    Lane 1 : Molecular weight marker
    Lane 2 : Cell lysates prepared from human Jurkat cells
    Lane 3 : Cell lysates prepared from human Jurkat cells treated with staurosporine
    Lane 4 : Cell lysates prepared from mouse NIH3T3 cells
    Lane 5 : Cell lysates prepared from CHO-K1 cells
    Lane 6 : Cell lysates prepared from Rat-2 cells
  • Overlay histogram showing HeLa cells stained with ab26350 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab26350, 1μg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was a mix of mouse IgG1 [ICIGG1], (ab91353, 1μg/1x106 cells), IgG2a [ICIGG2A], (ab91361, 1μg/1x106 cells), IgG2b [PLPV219], (ab91366, 1μg/1x106 cells), IgG3 [MG3-35], (ab18394, 1μg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • Ab26350 staining H2A.X  in Human placenta tissue sections by Immunohistochemistry (IHC-P - formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 5% BSA for 30 minutes at 22°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/100 in TBS) for 16 hours at 4°C. A diluted Biotin conjugated Goat anti-mouse polyclonal (1/200) was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-gamma H2A.X (phospho S139) antibody [9F3] (ab26350) at 1/1000 dilution

    Lane 1 : Untreated Zebrafish embryos.
    Lane 2 : Zebrafish embryos treated with 25 Gy of X-Ray.
    Lane 3 : Zebrafish embryos treated with 50 Gy of X-Ray

    All lanes : Goat anti-mouse HRP at 1/10000 dilution

    Please see abreview for Western blot details.

    See Abreview

  • ab26350 staining gamma H2A.X in pig spermatocytes by Immunocytochemistry/ Immunofluorescence. Cells were fixed with formaldehyde, permeabilized with Triton x100 and blocking with 0.15% BSA was performed for 30 minutes at 250C was performed. Samples were incubated with primary antibody (1/100: in PBS + 0.15% BSA+0.1% Tween 20) for 12 hours at 25°C. An Alexa Fluor®488-conjugated goat monoclonal to mouse IgG was used at dilution at 1/100 as secondary antibody.

    See Abreview


This product has been referenced in:
  • Song X  et al. Autophagy deficient keratinocytes display increased DNA damage, senescence and aberrant lipid composition after oxidative stress in vitro and in vivo. Redox Biol 11:219-230 (2017). ICC/IF ; Human . Read more (PubMed: 28012437) »
  • Xie Q  et al. Folate deficiency facilitates recruitment of upstream binding factor to hot spots of DNA double-strand breaks of rRNA genes and promotes its transcription. Nucleic Acids Res 45:2472-2489 (2017). Read more (PubMed: 27924000) »

See all 47 Publications for this product

Customer reviews and Q&As

Es freut mich zu hören, dass Sie unseren Antikörper ab26350 in Gefrierschnitten (IHC-Fr) testen möchten. Sie können den Gutscheincode zum Kauf eines weiteren Antikörpers einsetzen, sobald Sie uns ein Abreview ...

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Thank you for your patience.

For ab26350:
The lab let me know that theyhave not specifically isotyped this antibody so they do not knowfor certain which subisotype the antibody is.We only know it is mouse IgG.

The according is...

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Thank you for your reply.

We have 2 mouse monoclonals: ab26350 (IgG, I will inquire about the subisotype with the lab) and ab22551 (IgG1). Both have been tested in ICC/IF with human samples.

I will let you know what I find out abou...

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Thank you for your enquiry.

I can confirm that this antibody has not yet been tested forH2AX detection in yeast.

I contacted the originator who did a BLAST against human H2AX protein sequence but there was no yeast found on the li...

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Thank you for contacting us. There will be a slight immunogen difference between each antibody. However, each recognizes the same portion of the targe,t gamma H2A.X (phospho S139), and should work in your experiments. Two of these antibodies are ra...

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DISCOUNT CODE: *** Expiration date: April 3, 2012 Thanks for your call today, and I'm pleased you would like to accept our offer and test ab26350 in IHC-Fr. This code will give you 1 free primary antibody before the expiration date. To redeem this offe...

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Thank you for contacting Abcam. Included in this email are the instructions for returning your item/s to Abcam. When you return the goods, please ensure that you follow the procedures as below: - Return in the original Abcam box with the cooler box and...

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I have conducted a sequence alignment between the immunogen of ab26350 and the zebrafish histone H2AX however, the alignment was very poor. There was only 5% homology therefore unfortunately this antibody is not suitable for use in zebrafish.
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