PRODUCT CODE: ET1602-2

Phospho-Histone H2A.X (S139) Recombinant Rabbit Monoclonal Antibody [SR33-09] (ET1602-2)

  • Recombinant

Applications

  • WB

  • IHC-P

REACTIVITY

  • Human

  • Mouse

  • Rat

Western blot analysis of Phospho-Histone H2A.X(S139) on different lysates. Proteins were transferred to a PVDF membrane and blocked with 5% BSA in PBS for 1 hour at room temperature. The primary antibody (ET1602-2, 1/500) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:5,000 dilution was used for 1 hour at room temperature.<br />
Positive control: <br />
Lane 1: HepG2 cell lysate–treated with etoposide <br />
Lane 2: HepG2 cell lysate–untreated
  • Western blot analysis of Phospho-Histone H2A.X(S139) on different lysates. Proteins were transferred to a PVDF membrane and blocked with 5% BSA in PBS for 1 hour at room temperature. The primary antibody (ET1602-2, 1/500) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:5,000 dilution was used for 1 hour at room temperature.<br />
Positive control: <br />
Lane 1: HepG2 cell lysate–treated with etoposide <br />
Lane 2: HepG2 cell lysate–untreated
  • Immunohistochemical analysis of paraffin-embedded rat brain tissue using anti-Phospho-Histone H2A.X(S139) antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-2, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
  • Immunohistochemical analysis of paraffin-embedded mouse testis tissue using anti-Phospho-Histone H2A.X(S139) antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-2, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
  • Immunohistochemical analysis of paraffin-embedded mouse brain tissue using anti-Phospho-Histone H2A.X(S139) antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-2, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
  • Immunohistochemical analysis of paraffin-embedded mouse small intestine tissue using anti-Phospho-Histone H2A.X(S139) antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-2, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Western blot analysis of Phospho-Histone H2A.X(S139) on different lysates. Proteins were transferred to a PVDF membrane and blocked with 5% BSA in PBS for 1 hour at room temperature. The primary antibody (ET1602-2, 1/500) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:5,000 dilution was used for 1 hour at room temperature.
Positive control:
Lane 1: HepG2 cell lysate–treated with etoposide
Lane 2: HepG2 cell lysate–untreated

Applications

  • WB

  • IHC-P

REACTIVITY

  • Human

  • Mouse

  • Rat

SPECIFICATIONS

Product Type

Recombinant Rabbit monoclonal primary

Product Name

Phospho-Histone H2A.X (S139) Recombinant Rabbit Monoclonal Antibody [SR33-09] (ET1602-2)

Immunogen

Synthetic phospho-peptide corresponding to residues surrounding ser139 of human histone h2a.x.

Host

Rabbit

Modification

Phospho

Modification Site

S139

Positive Control

HepG2 cell lysate–treated with etoposide, HepG2 cell lysate, rat brain tissue, mouse testis tissue, mouse brain tissue, mouse small intestine tissue.

Conjugation

Unconjugated

Clonality

Monoclonal

Clone Number

SR33-09

PROPERTIES

Form

Liquid

Storage Condition

Store at +4C after thawing. Aliquot store at -20C or -80C. Avoid repeated freeze / thaw cycles.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Concentration

1 ug/ul

PURIFICATION

Protein A affinity purified.

MOLECULAR WEIGHT

15 kDa

Isotype

IgG

APPLICATION DILUTION

  • WB

  • 1:500-1:5,000

  • IHC-P

  • 1:50-1:200

TARGET

UNIPROT #

PROTEIN NAME

Phospho-Histone H2A.X (S139)

SYNONYMS

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; Histone H2AX antibody; RGD1566119 antibody

SEQUENCE SIMILARITIES

Belongs to the histone H2A family.

DEVELOPMENTAL STAGE

Synthesized in G1 as well as in S-phase.

POST-TRANSLATIONAL MODIFICATION

Phosphorylated on Ser-140 (to form gamma-H2AX 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 (By similarity). 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. Ubiquitination at Lys-14 and Lys-16 (H2AK13Ub and H2AK15Ub, respectively) in response to DNA damage is initiated by RNF168 that mediates monoubiquitination at these 2 sites, and 'Lys-63'-linked ubiquitin are then conjugated to monoubiquitin; RNF8 is able to extend 'Lys-63'-linked ubiquitin chains in vitro. H2AK119Ub and ionizing radiation-induced 'Lys-63'-linked ubiquitination (H2AK13Ub and H2AK15Ub) are distinct events.; Acetylation at Lys-37 increases in S and G2 phases. This modification has been proposed to play a role in DNA double-strand break repair (By similarity).

SUBCELLULAR LOCATION

Nucleus, Chromosome

FUNCTION

Histone H2A.X is a member of the Histone H2A family, which is involved in nucleosomal organization of chromatin. The H2AFX gene is located in close proximity to the Porphobilinogen deaminase (PBG-D) gene in both mouse and human, and maps to chromosome 9 and 11q23, respectively. H2A.X differs from the other members of the H2A family by the presence of a highly conserved C-terminal motif. It is rapidly phosphorylated in response to ionizing radiation and plays an important role in the recognition and repair of DNA double stranded breaks. The phosphorylated form of H2A.X, designated ?-H2A.X, forms nuclear foci at the heavy chain constant region of cells involved in class switch recombination (CSR), a region-specific DNA reaction that replaces one immunoglobulin heavy chain constant region gene with another. The phosphorylated ?-H2A.X is also thought to initiate subsequent repair factors, including Rad50, Rad51 and BRCA1.