Anti-Histone H4 (tri methyl K20)抗体- ChIP Grade (ab9053)

概述

  • 产品名称
    Anti-Histone H4 (tri 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 (tri methyl K20) - ChIP Grade
  • 特异性
    Histone H4 tri-methylated at Lys 20.
  • 经测试应用
    适用于: ChIP/Chip, IHC-P, Electron Microscopy, IHC (PFA fixed), IHC-Fr, ICC/IF, Flow Cyt, WB, ChIPmore details
  • 种属反应性
    与反应: Mouse, Rat, Cow, Human, Schizosaccharomyces pombe, Toxoplasma gondii, Xenopus tropicalis
    预测可用于: Mammal
  • 免疫原

    Synthetic peptide within Human Histone H4 aa 1-100 (tri methyl K20) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary.
    (Peptide available as ab17567)

  • 阳性对照
    • WB: Calf thymus histone preparation and HeLa whole cell extract. IHC-P: Human normal skin tissue.
  • 常规说明

    In immunofluorescence: Human cells: The antibody stains a subset of cellular chromatin that is characteristically DAPI-rich (condensed, heterochromatic). The perinucleolar heterochromatin is particularly rich in trimethylated lysine 20 staining. Mouse: Prominent staining of a subset of centromeric or pericentromeric heterochromatin. Interphase: The cells within the culture show a considerable variability in the intensity of staining with the antibody. The relationship between trimethylation levels and cell cycle have not yet been determined but may be a contributor to the amount of methylation detected in each cell. Heterochromatic regions of the interphase nucleus are the primary sites of trimethylation observed by indirect immunofluroescence. Mitosis: Discrete chromosomal regions are labelled intensely, with lower level fluorescence throughout the remainder of the chromosome arms. Immunofluorescence staining was performed as part of the nuclear antibody characterisation program at www.cellnucleus.com. Trimenthylation at Lys20 of Histone H4 isa common hallmark of human cancer (Fraga et al. 2005).

性能

  • 形式
    Liquid
  • 存放说明
    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.
  • 存储溶液
    Preservative: 0.02% Sodium Azide
    Constituents: 1% BSA, PBS, pH 7.4
  • Concentration information loading...
  • 纯度
    Immunogen affinity purified
  • 纯化说明
    Purified using Sulpholink column with specific peptide linked via its cysteine residue
  • Primary antibody说明
    In immunofluorescence: Human cells: The antibody stains a subset of cellular chromatin that is characteristically DAPI-rich (condensed, heterochromatic). The perinucleolar heterochromatin is particularly rich in trimethylated lysine 20 staining. Mouse: Prominent staining of a subset of centromeric or pericentromeric heterochromatin. Interphase: The cells within the culture show a considerable variability in the intensity of staining with the antibody. The relationship between trimethylation levels and cell cycle have not yet been determined but may be a contributor to the amount of methylation detected in each cell. Heterochromatic regions of the interphase nucleus are the primary sites of trimethylation observed by indirect immunofluroescence. Mitosis: Discrete chromosomal regions are labelled intensely, with lower level fluorescence throughout the remainder of the chromosome arms. Immunofluorescence staining was performed as part of the nuclear antibody characterisation program at www.cellnucleus.com. Trimenthylation at Lys20 of Histone H4 isa common hallmark of human cancer (Fraga et al. 2005).
  • 克隆
    多克隆
  • 同种型
    IgG
  • 研究领域

应用

Our Abpromise guarantee covers the use of ab9053 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
ChIP/Chip Use at an assay dependent concentration. PubMed: 19077033
IHC-P Use at an assay dependent concentration. PubMed: 20007528
Electron Microscopy Use at an assay dependent concentration. PubMed: 20543957
IHC (PFA fixed) Use at an assay dependent concentration. PubMed: 17083276
IHC-Fr Use at an assay dependent concentration. PubMed: 17712411
ICC/IF Use at an assay dependent concentration. PubMed: 19727073
Flow Cyt 1/100.

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

WB Use a concentration of 1 µg/ml. Detects a band of approximately 13 kDa (predicted molecular weight: 11 kDa).Can be blocked with Human Histone H4 (tri methyl K20) peptide (ab17567).
ChIP Use 4-5µg for 106 cells.

靶标

图片

  • Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 min. The  ChIP was performed with 25 µg of chromatin, 6 µl of  ab9053 (blue), and 20 µl 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). Primers and probes are located in the first kb of the transcribed region.

  • Immunohistochemical staining of paraffin embedded, PFA fixed, mouse liver using ab9053 at 1/100. Heat mediated antigen retrieval was performed using sodium citrate buffer, pH 6, and the sample was permeabilized with 0.05% Triton X 100. The sample was blocked in 5% BSA for 1 hour at room temperature and was then incubated with primary antibody for 3 hours in PBS/0.05% Triton X 100 at room temperature. An Alexa Fluor® 555 donkey anyti-rabbit was used as the secondary at 1/250.

    See Abreview



  • Predicted band size : 11 kDa

    ab9053 is specific for Histone H4 (tri-methyl K20). This is illustrated in lane 5 where the activity of ab9053 is specifically blocked by the addition of the immunizing peptide (ab17567).

  • A field of HeLa cells stained with anti-Trimethy Lysine 20 of Histone H4.

    This image is part of the nuclear antibody characterisation program at www.cellnucleus.com

  • IHC image of Histone H4 (tri methyl K20) staining in Human normal skin 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 ab9053, 0.2µg/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.

  • HeLa cells immunofluorescently stained with anti-Tri Methyl Lysine 20 of histone H4 (ab9053). HeLa cells were cultured on glass coverslips, fixed with 4% paraformaldehyde, and immunofluorescently labeled with anti-Tri K20 (green) at 1/500 and DAPI (red).

    This image is part of the nuclear antibody characterisation program at www.cellnucleus.com

    The image was collected using a 1.3 N.A. 40 X PlanApo Objective (Zeiss) using a Zeiss Axiovert 200M inverted fluorescent microscope and a SensiCam HQ (Cooke Corp.) 12-bit CCD. The images were collected using Universal Imaging MetaMorph v. 5.1 for Windows. A Physik Instruments piezo z-drive was used to collect images at 200 nm intervals throughout the depth of the cell nucleus. The digital images were processed to remove fluorescent background and colour composites were made from single focal planes using MetaMorph

  • A mouse 10T1/2 embryonic fibroblast immunofluorescently stained with anti Tri Methyl Lysine 20 of histone H4. Murine 10T1/2 embryonic fibroblasts were cultured on glass coverslips, fixed with paraformaldehyde and then immunofluorescently labeled using an anti-Tri Methyl K20 of histone H4 antibody at 1/500 (green). The nuclear chromatin was simultaneously visualized using DAPI (red).

    This image is part of the nuclear antibody characterisation program at www.cellnucleus.com

    The image was collected using a 1.4 N.A. 100 X PlanApo Objective (Zeiss) using a Zeiss Axiovert 200M inverted fluorescent microscope and a Photometrics Cascade 16-bit CCD. The images were collected using Universal Imaging MetaMorph v. 5.1 for Windows. A Physik Instruments piezo z-drive was used to collect images at 200 nm intervals throughout the depth of the cell nucleus. The digital images were processed to remove fluorescent background and colour com



  • Predicted band size : 11 kDa

    Left panel: a fission yeast whole cell extract and recombinant (r) fission yeast histone H4.

    Right panel: fission yeast whole cell extracts from strains with only a single wt (h4) or mutant (h4K20R) gene were immunoblotted.

    Review by Steven Sanders submitted 13 August 2004

  • All lanes : Anti-Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053) at 1 µg/ml (Calf thymus histone lysate)

    Lane 1 : Human Histone H4 peptide (ab2622)
    Lane 2 : Human Histone H4 peptide (ab2622)
    Lane 3 : Human Histone H4 (mono methyl K20) peptide (ab17043)
    Lane 4 : Human Histone H4 (di methyl K20) peptide (ab14964)
    Lane 5 : Human Histone H4 (tri methyl K20) peptide (ab17567)
    Lane 6 : Human Histone H3 (tri methyl K4) peptide (ab1342)
    Lane 7 : Human Histone H3 (tri methyl K9) peptide (ab1773)

    Blocking peptides at 1 µg/ml per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution

    Predicted band size : 11 kDa

文献

This product has been referenced in:
  • Asensio-Juan E  et al. The histone demethylase PHF8 is a molecular safeguard of the IFN? response. Nucleic Acids Res 45:3800-3811 (2017). ChIP ; Human . Read more (PubMed: 28100697) »
  • Wang X  et al. A covalently bound inhibitor triggers EZH2 degradation through CHIP-mediated ubiquitination. EMBO J 36:1243-1260 (2017). Read more (PubMed: 28320739) »

See all 118 Publications for this product

客户评价及客户问答

Application
Western blot
Sample
Saccharomyces cerevisiae Cell lysate - whole cell (Whole cell lysate)
Gel Running Conditions
Reduced Denaturing (15%)
Loading amount
60000 cells
Specification
Whole cell lysate
Blocking step
Odyssey blocking buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 50% · Temperature: 20°C
Username

Abcam user community

Verified customer

提交于 Aug 03 2016

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
Username

Abcam user community

Verified customer

提交于 Oct 07 2014

Application
Western blot
Loading amount
240000 cells
Gel Running Conditions
Reduced Denaturing (10%)
Sample
Mouse Cell lysate - whole cell (Primary Neural cells)
Specification
Primary Neural cells
Treatment
Supplemented with and without serum in the culture media.
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 28°C
Username

Abcam user community

Verified customer

提交于 Sep 12 2014

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Sample
Human Cell (HEK293)
Specification
HEK293
Permeabilization
Yes - triton
Fixative
Methanol
Username

Abcam user community

Verified customer

提交于 Apr 23 2014

Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (Human Embryonic kidney cell lines)
Specification
Human Embryonic kidney cell lines
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 Nov 25 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 12 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - whole (Striatum (Brain))
Loading amount
50 µg
Specification
Striatum (Brain)
Gel Running Conditions
Reduced Denaturing (12%)
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 Nov 07 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (Mouse ES cells)
Loading amount
50 µg
Specification
Mouse ES cells
Gel Running Conditions
Reduced Denaturing (15% SDS-PAGE)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: RT°C
Username

Abcam user community

Verified customer

提交于 Nov 11 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Hela Cells)
Loading amount
50 µg
Specification
Hela Cells
Gel Running Conditions
Reduced Denaturing (15% SDS-PAGE)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: RT°C
Username

Abcam user community

Verified customer

提交于 Nov 11 2011

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Schistosoma mansoni Purified protein (1 adult worm)
Loading amount
1000 cells
Specification
1 adult worm
Gel Running Conditions
Non-reduced Non-Denaturing (Native) (15% SDS page)
Blocking step
Milk as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C°C
Username

Mrs. Celine Cosseau

Verified customer

提交于 May 31 2011

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