Diagenode

H3K4me3 monoclonal antibody - Classic

Histone-Deacetylase-polyclonal-antibody-diagenode
Catalog Number
Format
Price
C15200152
(MAb-152-050)
50 µg/50 µl
$295.00
  Bulk order

Monoclonal antibody raised in mouse against histone H3, trimethylated at lysine 4 (H3K4me3), using a KLH-conjugated synthetic peptide.

Lot001-11
Concentration1.0 µg/µl
Species reactivityHuman, mouse, Nematodes, Arabidopsis
TypeMonoclonal
PurityProtein A/G purified
HostRabbit
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP/ChIP-seq * 2 μg/ChIP Fig 1, 2
ELISA 1:5,000 Fig 3
Western Blotting 1:1,000 Fig 4
Immunofluorescence 1:500 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

    ChIP

    Figure 1. ChIP results obtained with the Diagenode monoclonal antibody directed against H3K4me3
    ChIP assays were performed using human HeLa cells, the Diagenode monoclonal antibody against H3K4me3 (Cat. No. C15200152) and optimized PCR primer pairs for qPCR. ChIP was performed with the “Auto Histone ChIP-seq” kit (Cat. No. C01010022), using sheared chromatin from 1 million cells on the SX-8G IP-Star automated system. 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. Quantitative PCR was performed with primers specific for the promoter of the constitutively expressed GAPDH and c-fos genes, used as positive controls, and for exon 2 of the inactive myoglobin (MB) gene and the Sat2 satellite repeat, used as negative controls. Figure 1 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.

    ChIP-seq

    ChIP-seq

    ChIP-seq

    ChIP-seq

    Figure 2. ChIP-seq results obtained with the Diagenode monoclonal antibody directed against H3K4me3
    ChIP was performed on sheared chromatin from 1 million HeLaS3 cells using 2 μg of the Diagenode antibody against H3K4me3 (Cat. No. C15200152) 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 36 bp tags were aligned to the human genome using the ELAND algorithm. Figure 2 shows the peak distribution along two 5 Mb regions of chromosome 3 and 5 (figure 2A and B, respectively) and in two 100 kb regions surrounding the GAPDH and c-fos positive control genes (figure 2C and D). These results clearly show an enrichment of the H3K4 trimethylation at the promoters of active genes.

    ELISA

    Figure 3. Cross reactivity of the Diagenode monoclonal antibody directed against H3K4me3
    To test the specificity an ELISA was performed using a serial dilution of the Diagenode monoclonal antibody against H3K4me3 (Cat. No. C15200152). The wells were coated with peptides containing the unmodified H3K4 as well as the mono-, di- and trimethylated H3K4 and the trimethylated H3K9. Figure 3 shows a high specificity of the antibody for the modification of interest.

    Western blot

    Figure 4. Western blot analysis using the Diagenode monoclonal antibody directed against H3K4me3
    Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode monoclonal antibody against H3K4me3 (Cat. No. C15200152) diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left.

    Immunofluorescence

    Figure 5. Immunofluorescence using the Diagenode monoclonal antibody directed against H3K4me3
    HeLa cells were stained with the Diagenode antibody against H3K4me3 (Cat. No. C15200152) 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 1:500 in blocking solution followed by an anti-mouse antibody conjugated to Alexa594. The middle panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.

  • Testimonials

    I have extensively used the antibodies against the histone modifications H3K4me3, H3k27me3, H3K9ac, H4k8ac and H3K18ac provided by Diagenode. The high level of specificity and selectivity of these antibodies in mouse brain samples, confirmed by using several negative and positive controls run in parallel with mouse brain tissue samples, ensured successful and reproducible results. I have been a Diagenode costumer for over one year now and I am extremely satisfied with the efficiency of the Bioruptor Pico for chromatin shearing as well as all of the ChIP materials (i.e., antibodies, blocking peptides, primer pairs for qPCR) provided by this company. Many thanks.

    Dr. Ermelinda Lomazzo, Institute of Physiological Chemistry, AG Prof. Beat Lutz. University Medical Center of the Johannes Gutenberg University Mainz, Germany
  • Applications
    ELISA
    Enzyme-linked immunosorbent assay. 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-seq (ab)
    Read more
    ChIP-qPCR (ab)
    Read more
  • Documents
    Datasheet H3K4me3 C15200152 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...
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    Epigenetic Antibodies Brochure BROCHURE
    More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e...
    Download
  • Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H3K4me3 monoclonal antibody - Classic (Diagenode Cat# C15200152 Lot# 001-11). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Methylation of the Sox9 and Oct4 promoters and its correlation with gene expression during testicular development in the laboratory mouse
    Pamnani M et al.
    Sox9 and Oct4 are two important regulatory factors involved in mammalian development. Sox9, a member of the group E Sox transcription factor family, has a crucial role in the development of the genitourinary system, while Oct4, commonly known as octamer binding transcription factor 4, belongs to class V of the trans...

    Germline organization in Strongyloides nematodes reveals alternative differentiation and regulation mechanisms.
    Kulkarni A et al.
    Nematodes of the genus Strongyloides are important parasites of vertebrates including man. Currently, little is known about their germline organization or reproductive biology and how this influences their parasitic life strategies. Here, we analyze the structure of the germline in several Strongyloides and closely ...

    Transcription termination and chimeric RNA formation controlled by Arabidopsis thaliana FPA.
    Duc C, Sherstnev A, Cole C, Barton GJ, Simpson GG
    Alternative cleavage and polyadenylation influence the coding and regulatory potential of mRNAs and where transcription termination occurs. Although widespread, few regulators of this process are known. The Arabidopsis thaliana protein FPA is a rare example of a trans-acting regulator of poly(A) site choice. Analysi...

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    Oct 3-Oct 8, 2016
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    Boston, USA
    Oct 3-Oct 4, 2016
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    Sep 27-Sep 30, 2016
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