Anti-Histone H2A.X 抗体 - ChIP Grade (ab11175)

製品の概要

  • 製品名Anti-Histone H2A.X antibody - ChIP Grade
    Histone H2A.X 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to Histone H2A.X - ChIP Grade
  • アプリケーション適用あり: IHC-P, WB, IP, ChIP, ICC/IF, IHC-Frmore details
  • 種交差性
    交差種: Mouse, Rat, Human, Monkey
    交差が予測される動物種: Rabbit, Rhesus monkey, Gorilla
  • 免疫原

    This information is considered to be commercially sensitive.

  • ポジティブ・コントロール
    • Tested with human HEK293, human G-361 and mouse embryonic fibroblast cells.
  • 特記事項The phosphorylated form of this Ab is known as gamma H2A.X, when phosphorylated at Ser 139.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab11175 in the following tested applications.

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

アプリケーション Abreviews 特記事項
IHC-P Use at an assay dependent concentration.
WB 1/5000 - 1/15000. Detects a band of approximately 15 kDa.
IP Use at 5-20 µg/mg of lysate.
ChIP Use 2 µg for 25 µg of chromatin. PubMed: 19380460
ICC/IF 1/2500. (see review). Use periodate-lysine-PFA fixative.
IHC-Fr 1/1000.

ターゲット情報

  • 機能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.
  • 配列類似性Belongs to the histone H2A family.
  • 発生段階Synthesized in G1 as well as in S-phase.
  • ドメインThe [ST]-Q motif constitutes a recognition sequence for kinases from the PI3/PI4-kinase family.
  • 翻訳後修飾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.
  • 細胞内局在Nucleus. Chromosome.
  • Information by UniProt
  • 参照データベース
  • 別名
    • 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
    see all

Anti-Histone H2A.X antibody - ChIP Grade 画像

  • Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab11175 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

  • ICC/IF image of ab11175 stained Hela cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab11175, 1µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • ab11175 staining Histone H2A.X in mouse liver tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and blocked with 5% BSA for 60 minutes at 25°C; antigen retrieval was by heat mediation in a Na-citrate pH6 buffer. Samples were incubated with primary antibody (1/100 in PBS/Triton) for 3 hours at 25°C. A Alexa Flour® 555-conjugated donkey anti-rabbit IgG polyclonal (1/250) was used as the secondary antibody.

    See Abreview

  • ab11175 staining Histone H2A.X in Mouse testis tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde, permeabilized with 0.1% Triton X-100 and blocked with 4% BSA for 2 hours at 37°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/100 in diluent) for 48 hours at 4°C. ab6564 Goat polyclonal anti-Rabbit IgG - H&L Cy5® (1/300) was used as the secondary antibody.

    Staining left to right:
    1) DAPI;
    2) Histone H2A.X;
    3) Merge

    See Abreview

  • ab11175 staining Histone H2A.X in murine aorta by Immunohistochemistry (Frozen sections).
    Tissue was fixed in acetone, permeabilized using 0.3% Triton, blocked with 10% serum for 30 minutes at 22°C and then incubated with ab11175 at a 1/1000 dilution for 16 hours at 4°C. The secondary used was ab6793, sheep polyclonal secondary antibody to rabbit IgG - H&L (TR), used at a 1/1000 dilution.

    See Abreview

  • Samples: Nuclear extract from human 293T, human G-361, and wild-type (+/+) or H2AX knockout (-/-) mouse embryonic fibroblasts. Antibody: ab11175 used at 0.1 mcg/ml. Detection: Chemiluminescence with an exposure time of 5 seconds.

    Samples: Nuclear extract from human 293T, human G-361, and wild-type (+/+) or H2AX knockout (-/-) mouse embryonic fibroblasts. Antibody: ab11175 used at 0.1 mcg/ml. Detection: Chemiluminescence with an exposure time of 5 seconds.

  • These images were kindly supplied as part of the review submitted by Geza-Fejes-Toth.
  • Anti-Histone H2A.X antibody - ChIP Grade (ab11175) at 1/2000 dilution + Rat C6 Cells, whole cell lysate at 25 µg

    Secondary
    HRP conjugated goat anti-rabbit antibody at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Observed band size : 16 kDa (why is the actual band size different from the predicted?)


    Exposure time : 1 second

    This image is courtesy of an anonymous Abreview

    See Abreview

Anti-Histone H2A.X antibody - ChIP Grade (ab11175) 使用論文

This product has been referenced in:
  • Piya S  et al. Atg7 suppression enhances chemotherapeutic agent sensitivity and overcomes stroma-mediated chemoresistance in acute myeloid leukemia. Blood 128:1260-9 (2016). Read more (PubMed: 27268264) »
  • Ošina K  et al. Effects of an Antimutagenic 1,4-Dihydropyridine AV-153 on Expression of Nitric Oxide Synthases and DNA Repair-related Enzymes and Genes in Kidneys of Rats with a Streptozotocin Model of Diabetes Mellitus. Basic Clin Pharmacol Toxicol 119:458-463 (2016). Read more (PubMed: 27163882) »

See all 35 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Frozen sections)
Sample Mouse Tissue sections (Aorta)
Specification Aorta
Fixative Acetone
Permeabilization Yes - 0.3% Triton
Blocking step Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 22°C
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投稿 Dec 20 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Cell lysate - whole cell (vascular smooth muscle cells)
Loading amount 100000 cells
Specification vascular smooth muscle cells
Gel Running Conditions Reduced Denaturing
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 May 14 2010

Application Western blot
Sample Human Cell lysate - whole cell (U2os)
Gel Running Conditions Reduced Denaturing
Loading amount 20 µg
Treatment 50 J/m2 UV and harvested after 1 hour
Specification U2os
Blocking step Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
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投稿 Aug 12 2016

Application Western blot
Sample Human Cell lysate - whole cell (HeLa)
Gel Running Conditions Reduced Denaturing
Loading amount 10 µg
Specification HeLa
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 Jul 01 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (esophageal tumor cells)
Gel Running Conditions Non-reduced Denaturing (12%SDS)
Loading amount 80 µg
Specification esophageal tumor cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
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Miss. Qingyuan Yang

Verified customer

投稿 Jan 04 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Cell lysate - whole cell (3T3MEF)
Gel Running Conditions Non-reduced Denaturing (12.5% acrylamide-Bis gel)
Loading amount 50000 cells
Treatment UV20J
Specification 3T3MEF
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: RT°C
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投稿 Dec 14 2015

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
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投稿 Oct 07 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step BSA as blocking agent for 20 minute(s) · Concentration: 5% · Temperature: RT°C
Sample Human Cell (T98G Human brain glioblastoma)
Specification T98G Human brain glioblastoma
Permeabilization Yes - 0.1% v/v Triton X-100 pH 7.4 for 5 min at RT
Fixative Paraformaldehyde
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Dr. Dimitra Kalamida

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投稿 Dec 06 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (15%)
Sample Rat Cell lysate - nuclear (vascular smooth muscle cells)
Specification vascular smooth muscle cells
Treatment 80 uM tert-butyl BHP
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 Jun 24 2013

Thank you for contacting us.

ab11175 and ab18255 were never used togteher in Sandwich ELISA. These however can be used in Indirect ELISA.

The sELISA application totally based on difference in epiotopes, the antibodies recognize mea...

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