RabMAb

Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade (ab81299)

Overview

  • Product nameAnti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade
    See all Histone H2A.X primary antibodies
  • Description
    Rabbit monoclonal [EP854(2)Y] to Histone H2A.X (phospho S139) - ChIP Grade
  • Tested applicationsWB, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    A synthetic phospho-peptide corresponding to residues surrounding serine 139 of Human Histone H2A.X protein was used as the immunogen.

  • Positive control
    • Jurkat cell lysate and Human kidney transitional cell carcinoma.
  • General notes

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated ‘PUR’ on our product labels.

    If you have any questions regarding this update, please contact our Scientific Support team.

    Produced using Abcam’s RabMAb® technology. RabMAb® technology is covered by the following U.S. Patents, No. 5,675,063 and/or 7,429,487.

    Alternative versions available:

    Anti-Histone H2A.X antibody (HRP) - ChIP Grade [EP854(2)Y] (ab195190)

     

Properties

Applications

Our Abpromise guarantee covers the use of ab81299 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
WB 1/5000 - 1/10000. Predicted molecular weight: 15 kDa.
IHC-P 1/50 - 1/100. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Target

  • FunctionVariant 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 similaritiesBelongs to the histone H2A family.
  • Developmental stageSynthesized in G1 as well as in S-phase.
  • DomainThe [ST]-Q motif constitutes a recognition sequence for kinases from the PI3/PI4-kinase family.
  • Post-translational
    modifications
    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 localizationNucleus. Chromosome.
  • Information by UniProt
  • Database links
  • Alternative names
    • AW228881 antibody
    • H2A histone family member X antibody
    • H2A.FX antibody
    • H2A.X antibody
    • H2A/X antibody
    • H2AFX antibody
    • H2AX antibody
    • H2AX histone antibody
    • H2AX_HUMAN antibody
    • Hist5.2ax antibody
    • Histone 2A antibody
    • Histone 2AX antibody
    • Histone H2A.x antibody
    • RGD1566119 antibody
    see all

Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade images

  • 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.

  • All lanes : Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade (ab81299) at 1/10000 dilution (purified)

    Lane 1 : Untreated HepG2 cell lysate
    Lane 2 : HepG2 cell lysate treated with etoposide

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP goat anti-rabbit (H+L) at 1/1000 dilution

    Predicted band size : 15 kDa
    Observed band size : 14 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • IHC-P image of γH2A.X staining on Mouse testis sections using unpurified ab81299 (1:5000). The sections were deparaffinized and subjected to heat mediated antigen retrieval using citric acid. The sections were blocked using 1% BSA for 10 mins at 21°C. ab81299 was diluted 1:5000 in TBS buffer (containing BSA and Azide) and sections were then incubated with unpurified ab81299 for 2 hours at 21°C. The secondary antibody used was Biotin conjugated Goat polyclonal to Rabbit IgG (1:250).

    See Abreview

  • IHC-P image of γH2A.X staining on Rat testis sections using unpurified ab81299 (1:5000). The sections were deparaffinized and subjected to heat mediated antigen retrieval using citric acid. The sections were blocked using 1% BSA for 10 mins at 21°C. ab81299 was diluted 1:5000 in TBS buffer (containing BSA and Azide) and sections were then incubated with unpurified ab81299 for 2 hours at 21°C. The secondary antibody used was Biotin conjugated Goat polyclonal to Rabbit IgG (1:250).

    See Abreview

  • 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.

  • All lanes : Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade (ab81299) at 1/5000 dilution (purified)

    Lane 1 : Untreated HepG2 cell lysate
    Lane 2 : HepG2 cell lysate treated with etoposide

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP goat anti-rabbit (H+L) at 1/1000 dilution

    Predicted band size : 15 kDa
    Observed band size : 14 kDa (why is the actual band size different from the predicted?)

    Blocking buffer: 5% NFDM/TBST

    Dilution buffer: 5% NFDM/TBST

  • All lanes : Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade (ab81299) at 1/500000 dilution (unpurified)

    Lane 1 : Jurkat cell lysates
    Lane 2 : Jurkat cell lysates treated with etoposide

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP Labelled goat anti-rabbit at 1/2000 dilution

    Predicted band size : 15 kDa
    Observed band size : 15 kDa
  • Immunohistochemical analysis of formalin/PFA-fixed paraffin-embedded human kidney transitional cell carcinoma using unpurified ab81299 at a dilution of 1/100.

References for Anti-Histone H2A.X (phospho S139) antibody [EP854(2)Y] - ChIP Grade (ab81299)

This product has been referenced in:
  • Xie C  et al. Expression of ?H2AX in various gastric pathologies and its association with Helicobacter pylori infection. Oncol Lett 7:159-163 (2014). IHC ; Human . Read more (PubMed: 24348841) »
  • Ha GH  et al. TACC3 deregulates the DNA damage response and confers sensitivity to radiation and PARP inhibition. Oncogene 0:N/A (2014). Human . Read more (PubMed: 24769898) »

See all 14 Publications for this product

Product Wall

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Na-citrate pH6
Sample Mouse Tissue sections (Liver, P5)
Specification Liver, P5
Permeabilization Yes - Triton 0,05%
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Oct 07 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step Serum as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Sample Human Cell (HEK293)
Specification HEK293
Permeabilization Yes - triton
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Apr 24 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Mouse Tissue sections (Testis)
Specification Testis
Fixative Formaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization No
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Username

Mr. Carl Hobbs

Verified customer

Submitted May 15 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step BSA as blocking agent for 15 minute(s) · Concentration: 0.1% · Temperature: 37°C
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: citrate
Sample Mouse Tissue sections (skin)
Specification skin
Permeabilization No
Fixative formalin
Username

Abcam user community

Verified customer

Submitted Jun 03 2014

Application Western blot
Loading amount 20 µg
Gel Running Conditions Non-reduced Denaturing (12% gel)
Sample Human Cell lysate - whole cell (HEK293)
Specification HEK293
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

Submitted Apr 25 2014

Application Western blot
Loading amount 10 µg
Gel Running Conditions Reduced Denaturing (4-12% gradient gel)
Sample Human Cell lysate - other (lymphocytes)
Specification lymphocytes
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 37°C
Username

Abcam user community

Verified customer

Submitted Nov 21 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Human Tissue sections (Testis)
Specification Testis
Fixative Formaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization No
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Username

Mr. Carl Hobbs

Verified customer

Submitted May 15 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Rat Tissue sections (Testis)
Specification Testis
Fixative Formaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citric acid
Permeabilization No
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Username

Mr. Carl Hobbs

Verified customer

Submitted May 15 2013

Thank you for your enquiry.

Could you confirm this is product number ab81299?

I have investigated our records for this antibody ab81299 with my colleagues and can confirm that regrettably there has been an error. This antibody has...

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"