Diagenode

H3K27me3 monoclonal antibody - Classic (sample size)

RFX5-polyclonal-antibody-diagenode
Catalog Number
Format
Price
C15200181-10
10 µg
$80.00
  Bulk order
Other format

Monoclonal antibody raised in mouse against histone H3 trimethylated at lysine 27 (H3K27me3), using a KLH-conjugated synthetic peptide.

Lot001-11
Concentration1.0 µg/µl
Species reactivityHuman, Nematodes
TypeMonoclonal
PurityProtein A purified
HostMouse
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP * 1-2 μg/ChIP Fig 1
ELISA 1:3,000 Fig 2
Western Blotting 1:1,000 Fig 3
Immunofluorescence 1:500 Fig 4
* 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 H3K27me3
    ChIP assays were performed using human HeLa cells, the Diagenode monoclonal antibody against H3K27me3 (Cat. No. MAb-181-050) and optimized PCR primer sets for qPCR. ChIP was performed with the “AutoHistone ChIP-seq” kit (Cat. No. AB-Auto02-A016) on sheared chromatin from 1 million cells. A titration of the antibody consisting of 1, 2, 5, and 10 μg per ChIP experiment was analysed. IgG (2 μg/IP) was used as negative IP control. QPCR was performed with primers for the promoters of the active genes c-fos (Cat. No. pp-1004-050) and GAPDH as negative controls, and for the coding regions of the inactive genes MYT1 and TSH2B (Cat. No. pp-1041-050) as positive 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 H3K27me3 is preferably present at inactive genes.

    ELISA

    Figure 2. Cross reactivity of the Diagenode monoclonal antibody directed against H3K27me3
    To test the specificity an ELISA was performed using a serial dilution of the Diagenode monoclonal antibody against H3K27me3 (Cat. No. MAb-181-050). The wells were coated with peptides containing the unmodified H3K27 region as well as the mono-, di- and trimethylated H3K27 and the trimethylated H3K9. Figure 2 shows a high specificity of the antibody for the peptide containing the modification of interest.

    Western Blot

    Figure 3. Western blot analysis using the Diagenode monoclonal antibody directed against H3K27me3
    Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode monoclonal antibody against H3K27me3 (Cat. No. MAb-181-050) 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 4. Immunofluorescence using the Diagenode monoclonal antibody directed against H3K27me3
    HeLa cells were stained with the Diagenode antibody against H3K27me3 (Cat. No. MAb-181-050) 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 H3K27me3 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.

  • Applications
    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
    ELISA
    Enzyme-linked immunosorbent assay. 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
  • Documents
    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...
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    Datasheet H3K27me3 MAb-181-050 DATASHEET
    Datasheet description
    Download
  • Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H3K27me3 monoclonal antibody - Classic (sample size) (Diagenode Cat# C15200181-10 Lot# 001-11). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Chemical Biology Approaches for Characterization of Epigenetic Regulators
    Barsyte-Lovejoy D, Szewczyk MM, Prinos P, Lima-Fernandes E, Ackloo S, Arrowsmith CH
    Chemical biology approaches are a powerful means to functionally characterize epigenetic regulators such as histone modifying enzymes. We outline experimental protocols and best practices for the cellular characterization and use of “chemical probes” that selectively inhibit protein methyltransferases, m...

    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 ...

    An Orally Bioavailable Chemical Probe of the Lysine Methyltransferases EZH2 and EZH1.
    Konze KD, Ma A, Li F, Barsyte-Lovejoy D, Parton T, Macnevin CJ, Liu F, Gao C, Huang XP, Kuznetsova E, Rougie M, Jiang A, Pattenden SG, Norris JL, James LI, Roth BL, Brown PJ, Frye SV, Arrowsmith CH, Hahn KM, Wang GG, Vedadi M, Jin J
    EZH2 or EZH1 is the catalytic subunit of the polycomb repressive complex 2 that catalyzes methylation of histone H3 lysine 27 (H3K27). The trimethylation of H3K27 (H3K27me3) is a transcriptionally repressive post-translational modification. Overexpression of EZH2 and hypertrimethylation of H3K27 have been implicated...

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