Anti-Histone H3 (di methyl K4)抗体- ChIP Grade (ab7766)

概述

  • 产品名称Anti-Histone H3 (di methyl K4)抗体- ChIP Grade
    参阅全部 Histone H3 一抗
  • 描述
    兔多克隆抗体to Histone H3 (di methyl K4) - ChIP Grade
  • 经测试应用适用于: ChIP, Dot blot, WB, ChIP/Chip, PepArr, IHC-Fr, ICC/IF, IHC-Pmore details
  • 种属反应性
    与反应: Mouse, Human, Pig, Saccharomyces cerevisiae, Tetrahymena, Drosophila melanogaster, Schizosaccharomyces pombe, Plasmodium falciparum, Marmoset (common), Candida albicans
    预测可用于: all Mammals
  • 免疫原

    Synthetic peptide within Human Histone H3 aa 1-100 (di methyl K4) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary.
    (Peptide available as ab7768, ab125304, ab7766)

  • 阳性对照
    • Calf Thymus Histone Preparation
  • 常规说明

    For detection of Histone H3 specifically methylated at position Lys 4. This antibody was used in a screen by Dover et al, (2002) to isolate yeast mutants that are unable to methylate Lysine 4. In immunofluorescence, this antibody detects foci in the nucleus that are non-colocalising with condensed chromatin. The perinuclear and perinucleolar heterochromatin are not stained with this antibody.

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...
  • 纯度Immunogen affinity purified
  • Primary antibody说明For detection of Histone H3 specifically methylated at position Lys 4. This antibody was used in a screen by Dover et al, (2002) to isolate yeast mutants that are unable to methylate Lysine 4. In immunofluorescence, this antibody detects foci in the nucleus that are non-colocalising with condensed chromatin. The perinuclear and perinucleolar heterochromatin are not stained with this antibody.
  • 克隆多克隆
  • 同种型IgG
  • 研究领域

应用

Our Abpromise guarantee covers the use of ab7766 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 Use 2 µg for 25 µg of chromatin.
Dot blot 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).Can be blocked with Human Histone H3 (di methyl K4) peptide (ab7768).
ChIP/Chip Use at an assay dependent concentration.
PepArr Use a concentration of 0.2 - 2 µg/ml.
IHC-Fr 1/1000.
ICC/IF 1/500.
IHC-P 1/800.

靶标

  • 功能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-translational modifications of histones, also called histone code, and nucleosome remodeling.
  • 序列相似性Belongs to the histone H3 family.
  • 发展阶段Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
  • 翻译后修饰Acetylation is generally linked to gene activation. Acetylation on Lys-10 (H3K9ac) impairs methylation at Arg-9 (H3R8me2s). Acetylation on Lys-19 (H3K18ac) and Lys-24 (H3K24ac) favors methylation at Arg-18 (H3R17me).
    Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
    Asymmetric dimethylation at Arg-18 (H3R17me2a) by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 (H3R8me2s) by PRMT5 is linked to gene repression. Asymmetric dimethylation at Arg-3 (H3R2me2a) by PRMT6 is linked to gene repression and is mutually exclusive with H3 Lys-5 methylation (H3K4me2 and H3K4me3). H3R2me2a is present at the 3' of genes regardless of their transcription state and is enriched on inactive promoters, while it is absent on active promoters.
    Methylation at Lys-5 (H3K4me), Lys-37 (H3K36me) and Lys-80 (H3K79me) are linked to gene activation. Methylation at Lys-5 (H3K4me) facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 (H3K79me) is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are linked to gene repression. Methylation at Lys-10 (H3K9me) is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 (H3S10ph) and acetylation of H3 and H4. Methylation at Lys-5 (H3K4me) and Lys-80 (H3K79me) require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are enriched in inactive X chromosome chromatin.
    Phosphorylated at Thr-4 (H3T3ph) by GSG2/haspin during prophase and dephosphorylated during anaphase. Phosphorylation at Ser-11 (H3S10ph) by AURKB is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at Ser-11 (H3S10ph) by RPS6KA4 and RPS6KA5 is important during interphase because it enables the transcription of genes following external stimulation, like mitogens, stress, growth factors or UV irradiation and result in the activation of genes, such as c-fos and c-jun. Phosphorylation at Ser-11 (H3S10ph), which is linked to gene activation, prevents methylation at Lys-10 (H3K9me) but facilitates acetylation of H3 and H4. Phosphorylation at Ser-11 (H3S10ph) by AURKB mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. Phosphorylation at Ser-11 (H3S10ph) is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylated at Ser-29 (H3S28ph) by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation. Phosphorylation at Thr-7 (H3T6ph) by PRKCBB is a specific tag for epigenetic transcriptional activation that prevents demethylation of Lys-5 (H3K4me) by LSD1/KDM1A. At centromeres, specifically phosphorylated at Thr-12 (H3T11ph) from prophase to early anaphase, by DAPK3 and PKN1. Phosphorylation at Thr-12 (H3T11ph) by PKN1 is a specific tag for epigenetic transcriptional activation that promotes demethylation of Lys-10 (H3K9me) by KDM4C/JMJD2C. Phosphorylation at Tyr-42 (H3Y41ph) by JAK2 promotes exclusion of CBX5 (HP1 alpha) from chromatin.
    Monoubiquitinated by RAG1 in lymphoid cells, monoubiquitination is required for V(D)J recombination (By similarity). Ubiquitinated by the CUL4-DDB-RBX1 complex in response to ultraviolet irradiation. This may weaken the interaction between histones and DNA and facilitate DNA accessibility to repair proteins.
  • 细胞定位Nucleus. Chromosome.
  • Information by UniProt
  • 数据库链接
  • 别名
    • HIST1 cluster, H3J antibody
    • Histone gene cluster 1, H3 histone family, member E antibody
    • Histone gene cluster 1, H3G antibody
    • H3 histone family, member J antibody
    • HIST1 cluster, H3E antibody
    • HIST1 cluster, H3I antibody
    • Histone gene cluster 1, H3C antibody
    • FLJ92264 antibody
    • H3 histone family, member A antibody
    • H3 histone family, member B antibody
    • H3 histone family, member C antibody
    • H3 histone family, member D antibody
    • H3 histone family, member F antibody
    • H3 histone family, member H antibody
    • H3 histone family, member I antibody
    • H3 histone family, member K antibody
    • H3 histone family, member L antibody
    • H3 histone family, member T antibody
    • H3 histone, family 3A antibody
    • H3.1 antibody
    • H3.3A antibody
    • H3/a antibody
    • H3/b antibody
    • H3/c antibody
    • H3/d antibody
    • h3/f antibody
    • H3/h antibody
    • H3/i antibody
    • H3/j antibody
    • H3/k antibody
    • H3/l antibody
    • H3/t antibody
    • H31_HUMAN antibody
    • H3F1K antibody
    • H3F3 antibody
    • H3F3A antibody
    • H3FA antibody
    • H3FB antibody
    • H3FC antibody
    • H3FD antibody
    • H3FF antibody
    • H3FH antibody
    • H3FI antibody
    • H3FJ antibody
    • H3FK antibody
    • H3FL antibody
    • HIST1 cluster, H3A antibody
    • HIST1 cluster, H3B antibody
    • HIST1 cluster, H3C antibody
    • HIST1 cluster, H3D antibody
    • HIST1 cluster, H3F antibody
    • HIST1 cluster, H3G antibody
    • HIST1 cluster, H3H antibody
    • HIST1H3A antibody
    • HIST1H3B antibody
    • HIST1H3C antibody
    • HIST1H3D antibody
    • HIST1H3E antibody
    • HIST1H3F antibody
    • HIST1H3G antibody
    • HIST1H3H antibody
    • HIST1H3I antibody
    • HIST1H3J antibody
    • HIST3H3 antibody
    • Histone 1, H3a antibody
    • Histone 1, H3b antibody
    • Histone 1, H3c antibody
    • Histone 1, H3d antibody
    • Histone 1, H3e antibody
    • Histone 1, H3f antibody
    • Histone 1, H3g antibody
    • Histone 1, H3h antibody
    • Histone 1, H3i antibody
    • Histone 1, H3j antibody
    • histone 3, H3 antibody
    • histone cluster 1 H3 family member a antibody
    • histone cluster 1 H3 family member b antibody
    • histone cluster 1 H3 family member c antibody
    • histone cluster 1 H3 family member d antibody
    • histone cluster 1 H3 family member e antibody
    • histone cluster 1 H3 family member f antibody
    • histone cluster 1 H3 family member g antibody
    • histone cluster 1 H3 family member h antibody
    • histone cluster 1 H3 family member i antibody
    • histone cluster 1 H3 family member j antibody
    • Histone cluster 1, H3a antibody
    • Histone cluster 1, H3b antibody
    • Histone cluster 1, H3c antibody
    • Histone cluster 1, H3d antibody
    • Histone cluster 1, H3e antibody
    • Histone cluster 1, H3f antibody
    • Histone cluster 1, H3g antibody
    • Histone cluster 1, H3i antibody
    • Histone cluster 1, H3j antibody
    • Histone gene cluster 1, H3 histone family, member A antibody
    • Histone gene cluster 1, H3 histone family, member B antibody
    • Histone gene cluster 1, H3 histone family, member C antibody
    • Histone gene cluster 1, H3 histone family, member D antibody
    • Histone gene cluster 1, H3 histone family, member F antibody
    • Histone gene cluster 1, H3 histone family, member G antibody
    • Histone gene cluster 1, H3 histone family, member H antibody
    • Histone gene cluster 1, H3 histone family, member I antibody
    • Histone gene cluster 1, H3 histone family, member J antibody
    • Histone gene cluster 1, H3A antibody
    • Histone gene cluster 1, H3B antibody
    • Histone gene cluster 1, H3D antibody
    • Histone gene cluster 1, H3E antibody
    • Histone gene cluster 1, H3F antibody
    • Histone gene cluster 1, H3H antibody
    • Histone gene cluster 1, H3I antibody
    • Histone gene cluster 1, H3J antibody
    • Histone H 3 antibody
    • Histone H3.1 antibody
    • histone H3.1t antibody
    • Histone H3.2 antibody
    • Histone H3/a antibody
    • Histone H3/b antibody
    • Histone H3/c antibody
    • Histone H3/d antibody
    • Histone H3/f antibody
    • Histone H3/h antibody
    • Histone H3/i antibody
    • Histone H3/j antibody
    • Histone H3/k antibody
    • Histone H3/l antibody
    • Histone H3/m antibody
    • Histone H3/o antibody
    see all

Anti-Histone H3 (di methyl K4) antibody - ChIP Grade 图像

  • 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 ab7766 (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.

  • Anti-Histone H3 (di methyl K4) antibody - ChIP Grade (ab7766) at 1 µg/ml + Calf Thymus Histone Preparation Nuclear Lysate (ab121) at 0.5 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 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?)
    Additional bands at : 14 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 4 minutes
  • ab7766 at 1/200 staining human U2OS (osteosarcoma) cells by ICC/IF. The cells were paraformaldehyde fixed and then stained with the antibody for 1 hour. A Cy2 ® conjugated donkey anti-rabbit antibody was used as the secondary (green). The image shows uniformal staining of the whole nucleus, with several specles found. The insert shows H3-di methyl K4 of tw2 cells only. DAPI nuclear staining is shown in blue.

    See Abreview

  • IHC-P image of Histone H3 (di methyl K4) staining with ab7766 on human renal cancer tissue sections. The sections were subjected to heat-mediated antigen retrieval using citrate buffer pH6.0. The sections were then blocked with 5% serum for 20 minutes at 20°C, before incubation with ab7766 (1/500 dilution) for 45 minutes at 20°C. The secondary was a HRP conjugated goat anti-rabbit polyclonal, used at a 1/400 dilution.

    See Abreview

  • All batches of ab7766 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 - di methyl K4 peptide (ab7768), indicating that this antibody specifically recognises the Histone H3 - di methyl K4 modification.

    1. ab1340 - Histone H3 - mono methyl K4
    2. ab1342 - Histone H3 - tri methyl K4
    3. ab1771 - Histone H3 - mono methyl K9
    4. ab1772 - Histone H3 - di methyl K9
    5. ab1773 - Histone H3 - tri methyl K9
    6. ab1780 - Histone H3 - mono methyl K27
    7. ab1781 - Histone H3 - di methyl K27
    8. ab1782 - Histone H3 - tri methyl K27
    9. ab7228 - Histone H3 - unmodified
    10. ab7768 - Histone H3 - di methyl K4

  • Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 15 kDa

    MCF7 cells were incubated at 37&degC for 24h with vehicle control (0 &microM) and different concentrations of tranylcypromine hydrochloride (ab120606). Increased expression of Histone 3 K4 di-methyl (ab7766) in MCF7 cells correlates with an increase in tranylcypromine hydrochloride concentration, as described in literature.

    Nuclear extracts were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 10&microg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 5% BSA before being incubated with ab7766 at 1 &microg /ml and ab1791 at 1 &microg /ml overnight at 4°C. Antibody binding was detected using an anti-rabbit HRP secondary antibody (ab97051) at 1/10000 dilution and visualised using ECL development solution.

     

  • Dimethylated lysine 4 (green) is found in several hundred small nuclear foci that do not colocalize with condensed regions of chromatin (DAPI stained, red).  The perinuclear and perinucleolar heterochromatin do not stain with this antibody.
  • ICC/IF image of ab7766 stained HeLa cells. The cells were 4% formaldehyde fixed (10 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 (ab7766, 0.1µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, a goat anti-rabbit DyLight® 488 (IgG; H+L) 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. This antibody also gave a positive result in 4% formaldehyde fixed (10 min) Hek293, HepG2 and MCF7 cells at 0.1µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 0.1µg/ml.

Anti-Histone H3 (di methyl K4) antibody - ChIP Grade (ab7766)参考文献

This product has been referenced in:
  • Bledau AS  et al. The H3K4 methyltransferase Setd1a is first required at the epiblast stage, whereas Setd1b becomes essential after gastrulation. Development 141:1022-35 (2014). Read more (PubMed: 24550110) »
  • Stimpson KM  et al. Nucleolar organization, ribosomal DNA array stability, and acrocentric chromosome integrity are linked to telomere function. PLoS One 9:e92432 (2014). ChIP ; Human . Read more (PubMed: 24662969) »

See all 104 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Human Cell lysate - whole cell (HeLa cells treated with IFNg)
Negative control An adjacent coding region from SMOX genes. Exons of actively transcribed genes are enriched in H3K4me3.
Specification HeLa cells treated with IFNg
Detection step Real-time PCR
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% formaldehyde on rocker
Positive control Primers for a predicted enhancer in Chr 20 (Cell Cycle,Volume 14, Issue 2, pg253-260 2015). Enhancers have higher deposition of mono-methyl and di-methyl H3K4
Username

Mr. Yupen Chung

Verified customer

提交于 Feb 10 2016

Application Western blot
Sample Human Cell lysate - nuclear (HeLa and HEK 293T nuclear lysate)
Gel Running Conditions Reduced Denaturing (15% SDS gel)
Loading amount 10 µg
Specification HeLa and HEK 293T nuclear lysate
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Mr. Yupen Chung

Verified customer

提交于 Jan 08 2016

Application Western blot
Sample Human Cell lysate - nuclear (U87MG Glioblastome cell line)
Gel Running Conditions Reduced Denaturing (15%)
Loading amount 1 µg
Specification U87MG Glioblastome cell line
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Dr. Martin Engel

Verified customer

提交于 Nov 02 2015

Application ChIP
Detection step Real-time PCR
Sample Zebrafish Cell lysate - nuclear (whole embryo shield stage, 6 hours p.f.)
Specification whole embryo shield stage, 6 hours p.f.
Negative control We used PCR for zgc15 and igsf genomic regions (see additional information for primer sequences) as a reference control. No H3K4 mono- and di- methylation was expected in these regions based on previously published microarray data
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% in PBST
Positive control ChIP sample was 22 times enriched over input control in hoxb3a and 8 times in tiparp genomic region (see additional information for primer sequences). 10-20x enrichment in H3K4 mono- and di- methylation was expected in these regions based on previously published microarray data.
Username

Mrs. Daria Onichtchouk

Verified customer

提交于 Apr 01 2015

Application Immunocytochemistry/ Immunofluorescence
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 25°C
Sample Human Cell (HeLa)
Specification HeLa
Permeabilization Yes - tween-20
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

提交于 Jan 12 2015

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 25°C
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: pH 6.0 citric acid
Sample Human Tissue sections (Human invasive breast carcinoma)
Specification Human invasive breast carcinoma
Permeabilization Yes - tween-20
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

提交于 Jan 09 2015

Application Western blot
Loading amount 15 µg
Gel Running Conditions Reduced Denaturing (12% SDS-PAGE)
Sample Mouse Cell lysate - whole cell (MEL)
Specification MEL
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Jan 08 2015

Application Western blot
Loading amount 15 µg
Gel Running Conditions Reduced Denaturing
Sample Human Cell lysate - nuclear (U2OS)
Specification U2OS
Blocking step Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

提交于 May 15 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (Cell lines)
Loading amount 5 µg
Specification Cell lines
Gel Running Conditions Reduced Denaturing
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Feb 01 2013


For ab32356 (http://www.abcam.com/histone-h3-di-methyl-k4-antibody-y47-chip-grade-ab32356.html):
Based on our western blot testing testing, ab32356 should specifically detect Histone H3 dimethylated on Lysine 4. The WB figure on the product da...

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