Diagenode

H3K27me2 polyclonal antibody - Classic (sample size)

RFX5-polyclonal-antibody-diagenode
Catalog Number
Format
Price
C15410046-10
(pAb-046-050)
10 µg
$80.00
  Bulk order
Other format

Polyclonal antibody raised in rabbit against the histone H3, dimethylated at lysine 27 (H3K27me2), using a KLH-conjugated synthetic peptide.

LotA1968-0024P
Concentration2.63 µg/µl
Species reactivityHuman, zebrafish
TypePolyclonal
PurityAffinity purified
HostRabbit
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP * 2 μg/ChIP Fig 1
ELISA 1:1,000 Fig 2
Dot Blotting 1:50,000 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 antibody directed against H3K27me2
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H3K27me2 (cat. No. C15410046) and optimized PCR primer sets for qPCR. ChIP was performed with the “Auto Histone ChIP-seq” kit (cat. No. AB-Auto02-A100) on the SX-8G IP-Star Compact automated system, using 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 promoter of the active GAPDH and EIF4A2 genes, used as negative controls, and for the promoter of the inactive HBB and the coding region of the inactive MYOD1 genes, used 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 H3K27me2 is preferably present at silent genes.

    ELISA

    Figure 2. Determination of the antibody titer
    To determine the titer of the antibody, an ELISA was performed using a serial dilution of the Diagenode antibody directed against H3K27me2 (cat. No. C15410046). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution (Figure 2), the titer of the antibody was estimated to be 1:480,000.

    Dot Blot analysis

    Figure 3. Cross reactivity tests using the Diagenode antibody directed against H3K27me2
    A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K27me2 (cat. No. C15410046) with peptides containing other modifications of histone H3 and H4 and the unmodified sequence. One hundred to 0.2 pmol of peptide containing the respective histone modification were spotted on a membrane. The antibody was used at a dilution of 1:50,000. Figure 3 shows a high specificity of the antibody for the modification of interest.

    Western blot

    Figure 4. Western blot analysis using the Diagenode antibody directed against H3K27me2
    Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode antibody against H3K27me2 (cat. No. C15410046) 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 antibody directed against H3K27me2
    HeLa cells were stained with the Diagenode antibody against H3K27me2 (cat. No. C15410046) 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 H3K27me2 antibody (left) diluted 1:500 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.

  • 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
    DB
    Dot blotting 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...
    Download
    Epigenetic Antibodies Brochure BROCHURE
    More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e...
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    Datasheet H3K27me2 pAb-046-050 DATASHEET
    Datasheet description
    Download
  • Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H3K27me2 polyclonal antibody - Classic (sample size) (Diagenode Cat# C15410046-10 Lot# A1968-0024P). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Role of Annexin gene and its regulation during zebrafish caudal fin regeneration
    Saxena S, Purushothaman S, Meghah V, Bhatti B, Poruri A, Meena Lakshmi MG, Sarath Babu N, Murthy CL, Mandal KK, Kumar A, Idris MM
    The molecular mechanism of epimorphic regeneration is elusive due to its complexity and limitation in mammals. Epigenetic regulatory mechanisms play a crucial role in development and regeneration. This investigation attempted to reveal the role of epigenetic regulatory mechanisms, such as histone H3 and H4 lysine ac...

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