Anti-Histone H3 (citrulline R2 + R8 + R17) 抗体 - ChIP Grade (ab5103)

製品の概要

  • 製品名
    Anti-Histone H3 (citrulline R2 + R8 + R17) antibody - ChIP Grade
    Histone H3 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to Histone H3 (citrulline R2 + R8 + R17) - ChIP Grade
  • 特異性
    ab5103 detects a 17 kDa band in single lane Western Blot. Peptide inhibition in Western Blot hasn't been processed. Modification specificity is determined by Peptide Array. ab5103 binds strongly to Histone H3 citrulline 2 + 8 + 17 peptide.
  • アプリケーション
    適用あり: ICC/IF, PepArr, IHC-Fr, Flow Cyt, ChIP/Chip, WB, ChIPmore details
  • 種交差性
    交差種: Mouse, Rat, Rabbit, Cow, Human, Monkey
    交差が予測される動物種: a wide range of other species
  • 免疫原

    Synthetic peptide corresponding to Human Histone H3 aa 1-100 (citrulline R2 + R8 + R17) conjugated to Keyhole Limpet Haemocyanin (KLH). Also SwissProt: P84243, Q71DI3, Q16695, Q6NXT2.
    Database link: P68431
    (Peptide available as ab32876)

  • ポジティブ・コントロール
    • This antibody gave a positive signal in HL60 Whole Cell Lysate - DMSO and Calcium Ionophore treated. In WB ab5103 only recognizes human or bovine histone H3 when PADI4 and calcium are added.

製品の特性

  • 製品の状態
    Liquid
  • 保存方法
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • バッファー
    pH: 7.40
    Preservative: 0.02% Sodium azide
    Constituent: PBS

    Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
  • Concentration information loading...
  • 精製度
    Immunogen affinity purified
  • ポリ/モノ
    ポリクローナル
  • アイソタイプ
    IgG
  • 研究分野

アプリケーション

Our Abpromise guarantee covers the use of ab5103 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
ICC/IF Use at an assay dependent concentration. PubMed: 20733033
PepArr Use a concentration of 0.2 - 0.02 µg/ml.
IHC-Fr Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

ChIP/Chip Use at an assay dependent concentration.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa). Abcam recommends using 3-5% milk as the blocking agent
ChIP Use at an assay dependent concentration.

ターゲット情報

  • 関連性
    Core component of nucleosome. 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- modifications of histones, also called histone code, and nucleosome remodeling. P68431 and Q71DI3 are expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation. Q16695 is expressed in tesicular cells. Q6NXT2 is specifically expressed in the seminiferous tubules of testis and this Hominid-specific H3.5/H3F3C preferentially colocalizes with euchromatin, and it is associated with actively transcribed genes. P84243 is a variant histone H3 which replaces conventional H3 in a wide range of nucleosomes in active genes. Constitutes the predominant form of histone H3 in non-dividing cells and is incorporated into chromatin independently of DNA synthesis. Deposited at sites of nucleosomal displacement throughout transcribed genes, suggesting that it represents an epigenetic imprint of transcriptionally active chromatin. Post-translational modification. Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
  • 細胞内局在
    Nuclear
  • 参照データベース
  • 別名
    • Cit Histone H3 antibody
    • Citrullated antibody
    • citrullin antibody
    • Citrullinated Histone H3 antibody
    • citrulline antibody
    • H3 3 like sequence MH921 antibody
    • H3 3A antibody
    • H3 histone antibody
    • H3 histone family member E pseudogene antibody
    • H3F3 antibody
    • HIST3H3 antibody
    • Histone H3 3 pseudogene antibody
    • Histone H3 Citrullated antibody
    • Histone H3.1 antibody
    • Histone H3.1t antibody
    • Histone H3.2 antibody
    • Histone H3.3 antibody
    • Histone H3.3C antibody
    • Histone H3.5 antibody
    see all

画像

  • All lanes : Anti-Histone H3 (citrulline R2 + R8 + R17) antibody - ChIP Grade (ab5103) at 0.2 µg/ml

    Lane 1 : HL60 whole cell lysate (negative control)
    Lane 2 : HL60 whole cell lysate + DMSO (solvent control)
    Lane 3 : HL60 whole cell lysate + DMSO + Calcium Ionophore (positive control)

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat anti Rabbit IR680 at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 15 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)

    Loading Control: GAPDH

    This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab5103 overnight at 4°C. Antibody binding was detected using Goat anti Rabbit IR680 secondary at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

     

  • ab5103 staining Histone H3 (citrulline 2 + 8 + 17) in Mouse bone marrow cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed in formaldehyde and permeabilized in 0.1% Triton X-100 prior to blocking in 5% Goat serum for 2 hours at 25°C. The primary antibody was diluted 1/250 in PBS and incubated with the sample for 12 hours at 4°C. The secondary antibody was Alexa Fluor® 488-conjugated Goat anti-Rabbit polyclonal, diluted 1/500.
    Nuclei were counterstained blue with DAPI.

    See Abreview

  • All batches of ab5103 are tested in Peptide Array against peptides to different Histone H3 modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H3 - citrulline 2 + 8 + 17 peptide (ab32876), indicating that this antibody specifically recognises the Histone H3 - citrulline 2 + 8 + 17 modifications.

    ab32876 - Histone H3 - citrulline 2 + 8 + 17

    ab17566 - Histone H3 - unmodified

  • Chromatin immunoprecipitation using ab5103 on the pS2 promoter. Times are after stimulation by estrogen (Ul).


  • Predicted band size : 15 kDa

    Cuthbert, G.L. et al, (2004) Cell 118, 545-553

    Rabbit polyclonal to Histone H3 (citrulline 2 + 8 + 17) used at 1/2000 dilution, after blocking with TBST 5% BSA. Purified histones run out with approximately 250 ng of each histone.

    Lanes 1-3 contain Histone H3 (250 ng per lane)
    Lane 1: PADI4 + Calcium
    Lane 2: H3 + PADI4
    Lane 3: H3 + PADI4 + Calcium

    Lanes 4-5 contain bulk histones (250 ng per lane)
    Lane 4: PADI4
    Lane 5: PADI4 + Calcium

    Lane 6: MCF7 cell extract
    Lane 7: MCF7 cell extract (HA-PADI4)

    Secondary antibody : anti-rabbit HRP from Sigma.
    In WB ab5103 only recognizes human or bovine histone H3 when PADI4 and calcium are added.

  • ICC/IF image of ab5103 stained human HeLa cells. The cells were PFA fixed (10 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab5103, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).

  • All lanes : Anti-Histone H3 (citrulline R2 + R8 + R17) antibody - ChIP Grade (ab5103) at 1 µg/ml

    Lane 1 : HL60 (Human Caucasian promyelocytic leukaemia) DMSO and Calcium Ionophore treated Whole Cell Lysate with with 5% BSA
    Lane 2 : HL60 (Human Caucasian promyelocytic leukaemia) DMSO and Calcium Ionophore treated Whole Cell Lysate with with 5% milk
    Lane 3 : HL60 (Human Caucasian promyelocytic leukaemia) DMSO and Calcium Ionophore treated Whole Cell Lysate with with 3% milk

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 15 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above .

     

    Secondary antibody - goat anti-rabbit HRP (ab97051)

参考文献

This product has been referenced in:
  • Kamioka Y  et al. Intravital Förster resonance energy transfer imaging reveals osteopontin-mediated polymorphonuclear leukocyte activation by tumor cell emboli. Cancer Sci 108:226-235 (2017). IHC-P ; Mouse . Read more (PubMed: 27960041) »
  • Thomas DC  et al. Eros is a novel transmembrane protein that controls the phagocyte respiratory burst and is essential for innate immunity. J Exp Med 214:1111-1128 (2017). ICC/IF . Read more (PubMed: 28351984) »

See all 79 Publications for this product

レビューと Q&A

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Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Cell (Lung)
Permeabilization
Yes - Triton 0.1%
Specification
Lung
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 37°C
Fixative
Zinc fixative
Username

Abcam user community

Verified customer

投稿 Jul 26 2017

Application
ELISA
Sample
Human Serum (EDTA Plasma Samples)
Specification
EDTA Plasma Samples
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 21°C
Type
Direct
Username

Herr Dr. Thomas Scherz

Verified customer

投稿 Apr 28 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (MCF-7, Hep G2)
Gel Running Conditions
Reduced Denaturing
Loading amount
25 µg
Specification
MCF-7, Hep G2
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
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投稿 Aug 19 2016

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Kidney)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10 mM Citrate
Permeabilization
No
Specification
Kidney
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C
Fixative
Paraformaldehyde
Username

Abcam user community

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投稿 Oct 06 2015

Application
Western blot
Sample
Mouse Tissue lysate - whole (Kidney)
Gel Running Conditions
Reduced Denaturing (4-12%)
Loading amount
20 µg
Specification
Kidney
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Dr. Wesley Konsavage

Verified customer

投稿 Oct 05 2015

Application
Western blot
Loading amount
15 µg
Gel Running Conditions
Reduced Denaturing
Sample
Mouse Cell lysate - whole cell (hepatocytes)
Specification
hepatocytes
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
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投稿 Dec 26 2014

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Skin)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: EDTA pH 9.0
Permeabilization
No
Specification
Skin
Fixative
Formaldehyde
Username

Abcam user community

Verified customer

投稿 Jun 01 2012

Application
Western blot
Sample
Mouse Cell lysate - nuclear (Primary neurons)
Loading amount
40 µg
Specification
Primary neurons
Gel Running Conditions
Reduced Denaturing (18)
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 18°C
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投稿 Apr 18 2011

Application
Western blot
Sample
Human Cell lysate - nuclear (Keratinocytes)
Loading amount
50 µg
Specification
Keratinocytes
Gel Running Conditions
Reduced Denaturing (18)
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 18°C
Username

Abcam user community

Verified customer

投稿 Apr 18 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Bone marrow WBCs)
Specification
Bone marrow WBCs
Fixative
Formaldehyde
Permeabilization
Yes - 0.1% Triton X-100
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Feb 16 2011

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

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