Anti-Histone H4 (di methyl K20)抗体- ChIP Grade (ab9052)

概述

  • 产品名称Anti-Histone H4 (di methyl K20)抗体- ChIP Grade
    参阅全部 Histone H4 一抗
  • 描述
    兔多克隆抗体to Histone H4 (di methyl K20) - ChIP Grade
  • 经测试应用适用于: IP, WB, ChIP, ICC/IF, IHC-Pmore details
  • 种属反应性
    与反应: Mouse, Cow, Human, Schizosaccharomyces pombe, Toxoplasma gondii
    预测可用于: Rat, a wide range of other species不与反应: Saccharomyces cerevisiae
  • 免疫原

    Synthetic peptide within Saccharomyces cerevisiae Histone H4 aa 1-100 (di methyl K20) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary.
    (Peptide available as ab26213)

  • 阳性对照
    • WB: Calf thymus histone preparation and HeLa whole cell extract. IHC-P: FFPE human pancreas adenocarcinoma tissue.

性能

应用

Our Abpromise guarantee covers the use of ab9052 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
IP Use at an assay dependent concentration.
WB 1/2000. Detects a band of approximately 11 kDa.
ChIP Use at an assay dependent concentration. PubMed: 20829797
ICC/IF Use a concentration of 1 µg/ml.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

靶标

Anti-Histone H4 (di methyl K20) antibody - ChIP Grade 图像

  • ICC/IF image of ab9052 stained HeLa cells. The cells were 100% methanol fixed (5 min) then permeabilised using 0.1% PBS-Triton and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to further permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab9052 at 1µg/ml overnight at +4°C. The secondary antibody (pseudo-colored green) was Alexa Fluor® 488 goat anti- rabbit (ab150081) IgG (H+L) preadsorbed, used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1h at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

  • All lanes : Anti-Histone H4 (di methyl K20) antibody - ChIP Grade (ab9052) at 1/2000 dilution

    Lane 1 : yH4 17-24 peptide
    Lane 2 : Unmodified yH4 17-24 peptide
    Lane 3 : Mono methyl K20 yH4 17-24 peptide
    Lane 4 : Di methyl K20 yH4 17-24 peptide
    Lane 5 : Tri methyl K20 yH4 17-24 peptide

    Blocking peptides at 1 µg/ml per lane.


    Performed under reducing conditions.

    This image is courtesy of Steve Sanders, Tony Kouzarides lab, University of Cambridge

  • All lanes : Upper blot - Histone H4 antibody (gift of A. Verreault) Lower blot - Rabbit polyclonal to Histone H4 di methyl K20 (ab9052) at 1/2000

    Lane 1 : S.cerevisiae extract
    Lane 2 : S.pombe extract
    Lane 3 : S.pombe rH4
    Lane 4 : Drosophila rH4
    Lane 5 : Histones (Roche)

    Di methylation at K20 is seen in S. pombe and in calf thymus (Roche). S. cerevisiae lacks K20 (di) methylation.

  • Histone H4 (di methyl K20) was immunoprecipitated using 0.5mg HeLa whole cell extract, 5µg of Rabbit polyclonal to Histone H4 (di methyl K20) and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab9052.
    Secondary: Clean blot (HRP conjugate) at 1/1000 dilution.
    Band: 14kDa: Histone H4 (di methyl K20).

  • IHC image of ab9052 staining in human pancreas adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. 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 ab9052, 1µg/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.

    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.

Anti-Histone H4 (di methyl K20) antibody - ChIP Grade (ab9052)参考文献

This product has been referenced in:
  • Svensson JP  et al. A nucleosome turnover map reveals that the stability of histone H4 Lys20 methylation depends on histone recycling in transcribed chromatin. Genome Res 25:872-83 (2015). ChIP ; Schizosaccharomyces pombe . Read more (PubMed: 25778913) »
  • Bodor DL  et al. The quantitative architecture of centromeric chromatin. Elife 3:e02137 (2014). WB . Read more (PubMed: 25027692) »

See all 17 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Sample Caenorhabditis elegans Cell (Gut cell & embryos)
Specification Gut cell & embryos
Fixative Paraformaldehyde
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Abcam user community

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提交于 Sep 10 2015

Application Western blot
Sample Human Cell lysate - other (HeLa, mouse ES)
Gel Running Conditions Reduced Denaturing (4-12 % SDS PAGE)
Loading amount 2 µg
Specification HeLa, mouse ES
Blocking step Milk as blocking agent for 18 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
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提交于 Aug 04 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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Abcam user community

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提交于 Oct 07 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (multiple myeloma cells)
Loading amount 50 µg
Specification multiple myeloma cells
Gel Running Conditions Reduced Denaturing (10-20% tris glycine)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
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Abcam user community

Verified customer

提交于 Nov 21 2012

Abcam has not validated the combination of species/application used in this Abreview.
Application Western blot
Sample Caenorhabditis elegans Tissue lysate - nuclear (C. elegans)
Loading amount 10 µg
Specification C. elegans
Treatment 0, 75Gy r-ray
Gel Running Conditions Reduced Denaturing (12%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Ms. Jinsun Ryu

Verified customer

提交于 Sep 12 2012

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