Anti-Histone H4 (symmetric di methyl R3)抗体- ChIP Grade (ab5823)

概述

  • 产品名称Anti-Histone H4 (symmetric di methyl R3)抗体- ChIP Grade
    参阅全部 Histone H4 一抗
  • 描述
    兔多克隆抗体to Histone H4 (symmetric di methyl R3) - ChIP Grade
  • 经测试应用适用于: IHC-P, PepArr, ChIP, ICC/IF, WBmore details
  • 种属反应性
    与反应: Mouse, Cow, Human, Saccharomyces cerevisiae, Xenopus laevis, Arabidopsis thaliana, Drosophila melanogaster, Tobacco
    预测可用于: Rat, Pig, Rainbow Trout, Corn
  • 免疫原

    Synthetic peptide within Histone H4 aa 1-100 (symmetric di methyl R3) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary.
    (Peptide available as ab14791)

  • 阳性对照
    • WB: Calf thymus histone preparation and HeLa whole cell extract. IHC-P: Human normal kidney FFPE tissue sections.

性能

应用

Our Abpromise guarantee covers the use of ab5823 in the following tested applications.

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

应用 Ab评论 说明
IHC-P 1/500. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
PepArr Use a concentration of 0.2 - 0.02 µg/ml.
ChIP Use 2-4 µg for 25 µg of chromatin.
ICC/IF 1/2000.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 14 kDa (predicted molecular weight: 11.5 kDa).Can be blocked with Human Histone H4 (symmetric di methyl R3) peptide (ab14791).

靶标

Anti-Histone H4 (symmetric di methyl R3) antibody - ChIP Grade 图像

  • Anti-Histone H4 (symmetric di methyl R3) antibody - ChIP Grade (ab5823) at 1 µg + Calf Thymus Histone Preparation Nuclear Lysate at 0.5 µg

    Secondary
    Anti-Rabbit IgG VHH Single Domain Antibody (HRP) (ab191866) at 1/50000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 11.5 kDa
    Observed band size : 14 kDa (why is the actual band size different from the predicted?)


    Exposure time : 3 minutes

    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 2% Bovine Serum Albumin before being incubated with ab5823 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.

  • Chromatin was prepared from Hela cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The  ChIP was performed with 25µg of chromatin, 2µg of  ab5823 (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.

  • Interphase HeLa cell immunofluorescently labelled with anti-dimethylArg3 (H2A + H4) antibody (ab5823, 1/2000) and a polyclonal goat anti-Rabbit secondary antibody conjugated to Cy3 (1/200). ab5823 shows a strong nuclear focal staining pattern. For more information please see Abreview.

    See Abreview


  • IHC image of ab5823 staining Histone H4 (symmetric di methyl R3) in human kidney formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab5823, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • All batches of ab5823 are tested in Peptide Array against peptides to different Histone H4 and H2A 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 H4 - symmetric di methyl R3 peptide (ab14791), indicating that this antibody specifically recognises the Histone H4 - symmetric di methyl R3 modification.

    ab14791 - Histone H4 - symmetric di methyl R3

    ab17416 - Histone H4 asymmetric di methyl R3

    ab15821 - Histone H4 - unmodified

    ab22399 - Histone H2A symmetric di methyl R3

    ab22398 - Histone H2A asymmetric di methyl R3

    ab13186 - Histone H2A - unmodified

Anti-Histone H4 (symmetric di methyl R3) antibody - ChIP Grade (ab5823)参考文献

This product has been referenced in:
  • Deng X  et al. Protein arginine methyltransferase 5 functions as an epigenetic activator of the androgen receptor to promote prostate cancer cell growth. Oncogene N/A:N/A (2016). WB, ChIP ; Human . Read more (PubMed: 27546619) »
  • Wang WL  et al. Phosphorylation and arginine methylation mark histone H2A prior to deposition during Xenopus laevis development. Epigenetics Chromatin 7:22 (2014). IP ; Xenopus laevis . Read more (PubMed: 25302076) »

See all 27 Publications for this product

Product Wall

Application ChIP
Detection step Real-time PCR
Sample Mouse Cell lysate - whole cell (embryonic stem cell)
Specification embryonic stem cell
Negative control IgG
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: formaldehyde (1%)
Positive control H3K27me3 and H3K4me3
Username

Mr. Dan Stummer

Verified customer

提交于 Apr 18 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step Serum as blocking agent for 20 minute(s) · Concentration: 3% · Temperature: 23°C
Sample Mouse Cell (Mouse embryonic fibroblasts)
Specification Mouse embryonic fibroblasts
Permeabilization Yes - 0.3% Triton X-100
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

提交于 Nov 28 2013

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (12)
Sample Human Cell lysate - whole cell (Human HeLa cells)
Specification Human HeLa cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

提交于 Nov 18 2013

Application Western blot
Loading amount 5 µg
Gel Running Conditions Reduced Denaturing (12)
Sample Fruit fly (Drosophila melanogaster) Tissue lysate - nuclear (Fruit fly embryo)
Specification Fruit fly embryo
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

提交于 Nov 18 2013

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 40 µg
Gel Running Conditions Reduced Denaturing (15%)
Sample Mouse Cell lysate - whole cell (Neuro2A- neuroblastoma)
Specification Neuro2A- neuroblastoma
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Jun 07 2013

Thank you for your enquiry.



I blasted the sequence and there is a 100% homology between the immunogen and the sequence for Histone H4 of Drosophila melanogaster.



Please let us know if you have further questions.

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Read More
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 (Mouse brain)
Specification Mouse brain
Fixative Paraformaldehyde
Antigen retrieval step None
Permeabilization Yes - Titon X-100
Blocking step Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 18°C
Username

Abcam user community

Verified customer

提交于 Apr 18 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (Primary Neurons)
Specification Primary Neurons
Fixative Acetone
Permeabilization No
Blocking step BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Username

Abcam user community

Verified customer

提交于 Apr 18 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Mouse Cell lysate - nuclear (primary neurons)
Specification primary neurons
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 3 minute(s) and 0 second(s)
Detection step Real-time PCR
Username

Abcam user community

Verified customer

提交于 Apr 18 2011

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