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H3K4me3 polyclonal antibody

ChIP-seq Grade

Catalog Number
Format
Price
C15410030
(pAb-030-050)
50 µg
$380.00
  Bulk order
Other format
C15410030-10
10 µg



Polyclonal antibody raised in rabbit against the region of histone H3 containing the trimethylated lysine 4 (H3K4me3), using a KLH-conjugated synthetic peptide.

Lot002
Concentration1.4 µg/µl
Species reactivityHuman, mouse, Arabidopsis: positive. Other species: not tested.
TypePolyclonal
PurityAffinity purified polyclonal antibody.
HostRabbit
Storage ConditionsStore at -20°C; for long storage, store at -80°C. Avoid multiple freeze-thaw cycles.
Storage BufferPBS containing 0.05% azide and 0.05% ProClin 300.
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP/ChIP-seq * 1 µg/ChIP Fig 1, 2
Dot Blotting 1:2,000 Fig 3
Western Blotting 1:500 Fig 4
Immunofluorescence 1:100 Fig 5

* Please note that the optimal antibody amount per IP should be determined by the end-user. We recommend testing 1-5 µg per IP.

  • Validation data

    A. H3K4me3 Antibody ChIP Grade

    B. H3K4me3 Antibody ChIP Grade

    Figure 1. ChIP results obtained with the Diagenode antibody directed against H3K4me3
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H3K4me3 (Cat. No. C15410030) and optimized PCR primer pairs for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (cat. No. C01010051), using sheared chromatin from 1 million cells. The chromatin was spiked with a panel of in vitro assembled nucleosomes, each containing a specific lysine methylation (SNAP-ChIP K-MetStat Panel, Epicypher). A titration consisting of 1, 2, 5 and 10 µg of antibody per ChIP experiment was analyzed. IgG (2 µg/IP) was used as a negative IP control. Figure 1A. Quantitative PCR was performed with primers specific for the promoter of the active GAPDH and EIF4A2 genes, used as positive controls, and for the inactive MYOD1 gene, used as negative control. The graph shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis). These results are in accordance with the observation that trimethylation of K4 at histone H3 is associated with the promoters of active genes Figure 1B. Recovery of the nucleosomes carrying the H3K4me1, H3K4me2, H3K4me3 modifications and the unmodified H3K4 as determined by qPCR. The figure clearly shows the antibody is very specific in ChIP for the H3K4me3 modification. At higher concentrations some H3K4me2 is also precipitated.

    A. H3K4me3 Antibody ChIP-seq Grade

    B. H3K4me3 Antibody for ChIP-seq

    C. H3K4me3 Antibody for ChIP-seq assay

    D. H3K4me3 Antibody validated in ChIP-seq

    Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against H3K4me3
    ChIP was performed on sheared chromatin from 1 million HeLa cells using 1 µg of the Diagenode antibody against H3K4me3 (Cat. No. C15410030) as described above. The IP'd DNA was subsequently analysed on an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the peak distribution along the complete sequence and a 1.2 Mb region of the X-chromosome (figure 2A and B) and in two regions surrounding the GAPDH and EIF4A2 positive control genes, respectively (figure 2C and D). These results clearly show an enrichment of the H3K4 trimethylation at the promoters of active genes.

    H3K4me3 Antibody validated in Dot Blot

    Figure 3. Cross reactivity test using the Diagenode antibody directed against H3K4me3
    A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K4me3 (Cat. No. C15410030) with peptides containing other modifications and unmodified sequences of histone H3 and H4. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:2,000. Figure 3 shows a high specificity of the antibody for the modification of interest.

    H3K4me3 Antibody validated in Western Blot

    Figure 4. Western blot analysis using the Diagenode antibody directed against H3K4me3
    Western blot was performed on whole cell (40 µg, lane 1) and histone extracts (15 µg, lane 2) from HeLa cells, and on 1 µg of recombinant histone H2A, H2B, H3.1, H3.3 and H4 (lane 3, 4, 5, 6 and 7, respectively) using the Diagenode antibody against H3K4me3 (Cat. No. C15410030). The antibody was diluted 1:500 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is shown on the right, the marker (in kDa) is shown on the left.

    H3K4me3 Antibody validated in Immunofluorescence

    Figure 5. Immunofluorescence using the Diagenode antibody directed against H3K4me3
    HeLa cells were stained with the Diagenode antibody against H3K4me3 (Cat. No. C15410030) and with DAPI. Cells were fixed with 4% formaldehyde for 10’ and blocked with PBS/TX-100 containing 5% normal goat serum and 1% BSA. The cells were immunofluorescently labelled with the H3K4me3 antibody (left) diluted 1:100 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa488. The middle panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.

  • Target Description

    Histones are the main constituents of the protein part of chromosomes of eukaryotic cells. They are rich in the amino acids arginine and lysine and have been greatly conserved during evolution. Histones pack the DNA into tight masses of chromatin. Two core histones of each class H2A, H2B, H3 and H4 assemble and are wrapped by 146 base pairs of DNA to form one octameric nucleosome. Histone tails undergo numerous post-translational modifications, which either directly or indirectly alter chromatin structure to facilitate transcriptional activation or repression or other nuclear processes. In addition to the genetic code, combinations of the different histone modifications reveal the so-called “histone code”. Histone methylation and demethylation is dynamically regulated by respectively histone methyl transferases and histone demethylases. Trimethylation of histone H3K4 is associated with active promoters.

  •  実験手法
    DB
    Dot blotting Read more
    WB
    Western blot : The quality of antibodies used in this technique is crucial for correct and specific protein identification. Diagenode offers huge selection of highly sensitive and specific western blot-validated antibodies. Learn more about: Load... Read more
    IF
    Immunofluorescence: Diagenode offers huge selection of highly sensitive antibodies validated in IF. Immunofluorescence using the Diagenode monoclonal antibody directed against CRISPR/Cas9 HeLa cells transfected with a Cas9 expression vector (... Read more
    ChIP-qPCR (ab)
    Read more
    ChIP-seq (ab)
    Read more
  •  資料
    Datasheet H3K4me3 C15410030 DATASHEET
    Datasheet description    		 Download
    Antibodies you can trust POSTER
    Epigenetic research tools have evolved over time from endpoint PCR to qPCR to the analyses of lar...    		 Download
    Epigenetic Antibodies Brochure BROCHURE
    More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e...    		 Download
  •  Safety sheets
    H3K4me3 antibody SDS GB en Download
    H3K4me3 antibody SDS US en Download
    H3K4me3 antibody SDS DE de Download
    H3K4me3 antibody SDS JP ja Download
    H3K4me3 antibody SDS BE nl Download
    H3K4me3 antibody SDS FR fr Download
    H3K4me3 antibody SDS BE fr Download
    H3K4me3 antibody SDS ES es Download
  •  出版物

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H3K4me3 polyclonal antibody (Diagenode Cat# C15410030 Lot# 002). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    SUMO protease FUG1, histone reader AL3 and the PRC1 Complex areintegral to repeat-expansion induced epigenetic silencing in Arabidopsisthaliana
    Sureshkumar S. et al.
    Epigenetic gene silencing induced by expanded repeats can cause diverse phenotypes ranging from severe growth defects in plants to genetic diseases such as Friedreich’s ataxia in humans1. The molecular mechanisms underlying repeat expansion-induced epigenetic silencing remain largely unknown2,3. Using a plant ...

    Temporal modification of H3K9/14ac and H3K4me3 histone marksmediates mechano-responsive gene expression during the accommodationprocess in poplar
    Ghosh R. et al.
    Plants can attenuate their molecular response to repetitive mechanical stimulation as a function of their mechanical history. For instance, a single bending of stem is sufficient to attenuate the gene expression in poplar plants to the subsequent mechanical stimulation, and the state of desensitization can last for ...

    DNA sequence and chromatin modifiers cooperate to confer epigeneticbistability at imprinting control regions.
    Butz S. et al.
    Genomic imprinting is regulated by parental-specific DNA methylation of imprinting control regions (ICRs). Despite an identical DNA sequence, ICRs can exist in two distinct epigenetic states that are memorized throughout unlimited cell divisions and reset during germline formation. Here, we systematically study the ...

    Altered Chromatin States Drive Cryptic Transcription in AgingMammalian Stem Cells.
    McCauley Brenna S et al.
    A repressive chromatin state featuring trimethylated lysine 36 on histone H3 (H3K36me3) and DNA methylation suppresses cryptic transcription in embryonic stem cells. Cryptic transcription is elevated with age in yeast and nematodes, and reducing it extends yeast lifespan, though whether this occurs in mammals is unk...

    Age-associated cryptic transcription in mammalian stem cells is linked topermissive chromatin at cryptic promoters
    McCauley B. S. et al.
    Suppressing spurious cryptic transcription by a repressive intragenic chromatin state featuring trimethylated lysine 36 on histone H3 (H3K36me3) and DNA methylation is critical for maintaining self-renewal capacity in mouse embryonic stem cells. In yeast and nematodes, such cryptic transcription is elevated with age...

    Identification of Novel Molecular Markers of Human Th17 Cells.
    Sałkowska A, Karaś K, Karwaciak I, Walczak-Drzewiecka A, Krawczyk M, Sobalska-Kwapis M, Dastych J, Ratajewski M
    Th17 cells are important players in host defense against pathogens such as , , and . Th17 cell-mediated inflammation, under certain conditions in which balance in the immune system is disrupted, is the underlying pathogenic mechanism of certain autoimmune disorders, e.g., rheumatoid arthritis, Graves' disease, multi...

    The Chromatin Factor HNI9 and ELONGATED HYPOCOTYL5 Maintain ROS Homeostasis under High Nitrogen Provision.
    Bellegarde F, Maghiaoui A, Boucherez J, Krouk G, Lejay L, Bach L, Gojon A, Martin A
    Reactive oxygen species (ROS) can accumulate in cells at excessive levels, leading to unbalanced redox states and to potential oxidative stress, which can have damaging effects on the molecular components of plant cells. Several environmental conditions have been described as causing an elevation of ROS production i...

    Polycomb Repressive Complex 2 attenuates the very high expression of the Arabidopsis gene NRT2.1.
    Bellegarde F, Herbert L, Séré D, Caillieux E, Boucherez J, Fizames C, Roudier F, Gojon A, Martin A
    PRC2 is a major regulator of gene expression in eukaryotes. It catalyzes the repressive chromatin mark H3K27me3, which leads to very low expression of target genes. NRT2.1, which encodes a key root nitrate transporter in Arabidopsis, is targeted by H3K27me3, but the function of PRC2 on NRT2.1 remains unclear. Here, ...

    Interplay of cell–cell contacts and RhoA/MRTF‐A signaling regulates cardiomyocyte identity
    Dorn et al
    Cell–cell and cell–matrix interactions guide organ development and homeostasis by controlling lineage specification and maintenance, but the underlying molecular principles are largely unknown. Here, we show that in human developing cardiomyocytes cell–cell contacts at the intercalated disk connect...

    Neonatal exposure to hyperoxia leads to persistent disturbances in pulmonary histone signatures associated with NOS3 and STAT3 in a mouse model.
    Chao CM, van den Bruck R, Lork S, Merkle J, Krampen L, Weil PP, Aydin M, Bellusci S, Jenke AC, Postberg J
    Background: Early pulmonary oxygen exposure is one of the most important factors implicated in the development of bronchopulmonary dysplasia (BPD). Methods: Here, we analyzed short- and long-term effects of neonatal hyperoxia on NOS3 and STAT3 expression and corresponding epigenetic signatures using a hyperoxia-base...

    Rapid Communication: The correlation between histone modifications and expression of key genes involved in accumulation of adipose tissue in the pig.
    Kociucka B. et al.
    Histone modification is a well-known epigenetic mechanism involved in regulation of gene expression; however, it has been poorly studied in adipose tissues of the pig. Understanding the molecular background of adipose tissue development and function is essential for improving production efficiency and meat quality. ...

    Lhx2 interacts with the NuRD complex and regulates cortical neuron subtype determinants Fezf2 and Sox11
    Muralidharan B. et al.
    n the developing cerebral cortex, sequential transcriptional programs take neuroepithelial cells from proliferating progenitors to differentiated neurons with unique molecular identities. The regulatory changes that occur in the chromatin of the progenitors are not well understood. During deep layer neurogenesis, we...

    Deep sequencing and de novo assembly of the mouse oocyte transcriptome define the contribution of transcription to the DNA methylation landscape
    Veselovska L, Smallwood SA, Saadeh H, Stewart KR, Krueger F, Maupetit-Méhouas S, Arnaud P, Tomizawa S, Andrews S, Kelsey G
    BACKGROUND: Previously, a role was demonstrated for transcription in the acquisition of DNA methylation at imprinted control regions in oocytes. Definition of the oocyte DNA methylome by whole genome approaches revealed that the majority of methylated CpG islands are intragenic and gene bodies are hypermethylated...

    CpG signalling, H2A.Z/H3 acetylation and microRNA-mediated deferred self-attenuation orchestrate foetal NOS3 expression.
    Postberg J, Kanders M, Forcob S, Willems R, Orth V, Hensel KO, Weil PP, Wirth S, Jenke AC
    BACKGROUND: An adverse intrauterine environment leads to permanent physiological changes including vascular tone regulation, potentially influencing the risk for adult vascular diseases. We therefore aimed to monitor responsive NOS3 expression in human umbilical artery endothelial cells (HUAEC) and to study the unde...

    Paclitaxel resistance increases oncolytic adenovirus efficacy via upregulated CAR expression and dysfunctional cell cycle control.
    Ingemarsdotter CK, Tookman LA, Browne A, Pirlo K, Cutts R, Chelela C, Khurrum KF, Leung EY, Dowson S, Webber L, Khan I, Ennis D, Syed N, Crook TR, Brenton JD, Lockley M, McNeish IA
    Resistance to paclitaxel chemotherapy frequently develops in ovarian cancer. Oncolytic adenoviruses are a novel therapy for human malignancies that are being evaluated in early phase trials. However, there are no reliable predictive biomarkers for oncolytic adenovirus activity in ovarian cancer. We investigated the ...

    p53-Independent regulation of p21Waf1/Cip1 expression and senescence by PRMT6.
    Phalke S, Mzoughi S, Bezzi M, Jennifer N, Mok WC, Low DH, Thike AA, Kuznetsov VA, Tan PH, Voorhoeve PM, Guccione E
    p21 is a potent cyclin-dependent kinase inhibitor that plays a role in promoting G1 cell cycle arrest and cellular senescence. Consistent with this role, p21 is a downstream target of several tumour suppressors and oncogenes, and it is downregulated in the majority of tumours, including breast cancer. Here, we repor...

    Transcription and histone methylation changes correlate with imprint acquisition in male germ cells.
    Henckel A, Chebli K, Kota SK, Arnaud P, Feil R
    Genomic imprinting in mammals is controlled by DNA methylation imprints that are acquired in the gametes, at essential sequence elements called 'imprinting control regions' (ICRs). What signals paternal imprint acquisition in male germ cells remains unknown. To address this question, we explored histone methylation ...

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