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H3K27me3 polyclonal antibody - Classic

Catalog Number
100 µl

As an alternative we offer the purified H3K27me3 polyclonal antibody - Classic.

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Concentrationnot determined
Species reactivityHuman
PurityWhole antiserum
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP 5-10 μl/ ChIP Fig 1
ELISA 1:500 Fig 2
Dot Blotting 1:20,000 Fig 3
Western Blotting 1:1,000 Fig 4
Immunofluorescence 1:200 Fig 5
  • Validation Data


    Figure 1. ChIP results obtained with the Diagenode antibody directed against H3K27me3
    ChIP assays were performed using HeLa cells, the Diagenode antibody against H3K27me3 (Cat. No. CS-069-100) and optimized primer sets for qPCR. ChIP was performed with the “OneDay ChIP” kit (Cat. No. kch-oneDIP-060), using sheared chromatin from 1.6 million cells. A titration consisting of 5, 10, and 15 μl of antibody per ChIP experiment was analyzed. IgG (5 μg/IP) was used as a negative IP control. PCR was performed with primers specific for the promoter of the constitutively expressed c-fos gene (Cat. No. pp-1004-050, used as a negative control target) and for the satellite repeat Sat2 (Cat. No. pp-1040-050, used as a positive control target). Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).


    Figure 2. Determination of the antibody titer
    To determine the titer of the antibody, an ELISA was performed using a serial dilution of Diagenode antibody directed against H3K27me3 (Cat. No. CS-069-100) in antigen coated wells. 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 1:24,700.

    Dot Blot

    Figure 3. Cross reactivity tests using the Diagenode antibody directed against H3K27me3
    A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K27me3 (Cat. No. CS-069-100) with peptides containing other histone modifications including mono- and dimethylation of the same lysine and mono-, di- and trimethylation of other lysines of histone H3. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. 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 against H3K27me3
    Histone extracts of HeLa cells (HeLa HE, 15 μg) were analysed by Western blot using the Diagenode antibody against H3K27me3 (Cat. No. CS-069-100) 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.

    Figure 5. Immunofluorescence using the Diagenode antibody directed against H3K27me3
    NIH3T3 mouse fibroblasts were stained with the Diagenode antibody against H3K27me3 (Cat. No. CS-069-100) and with DAPI. Cells were formaldehyde fixed, permeabilized with Triton X100 and then blocked with PBS containing 1% BSA (Figure 5). (A) Cells were immunofluorescently labelled with the H3K27me3 antibody (diluted 1:200 and incubated for 1 hour at room temperature) followed by goat anti-rabbit antibody conjugated to FITC. (B) Staining of the nuclei with DAPI, which specifically labels DNA. Both antibody and DAPI staining are restricted to the nucleus. H3K27me3 shows a characteristic broadly dispersed pattern in interphase chromatin.

  •  Applications
    Enzyme-linked immunosorbent assay. Read more
    Dot blotting Read more
    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
    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
    Datasheet H3K27me3 CS-069-100 DATASHEET
    Datasheet description
  •  Publications

    How to properly cite this product in your work

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    Ordered chromatin changes and human X chromosome reactivation by cell fusion-mediated pluripotent reprogramming
    Cantone I et al.
    Erasure of epigenetic memory is required to convert somatic cells towards pluripotency. Reactivation of the inactive X chromosome (Xi) has been used to model epigenetic reprogramming in mouse, but human studies are hampered by Xi epigenetic instability and difficulties in tracking partially reprogrammed iPSCs. Here ...

    Spatiotemporal control of estrogen-responsive transcription in ERα-positive breast cancer cells.
    P-Y Hsu, H-K Hsu, T-H Hsiao, Z Ye, E Wang, A L Profit, I Jatoi, Y Chen, N B Kirma, V X Jin, Z D Sharp and T H-M Huang
    Recruitment of transcription machinery to target promoters for aberrant gene expression has been well studied, but underlying control directed by distant-acting enhancers remains unclear in cancer development. Our previous study demonstrated that distant estrogen response elements (DEREs) located on chromosome 20q13...

    Epstein-Barr virus-mediated transformation of B cells induces global chromatin changes independent to the acquisition of proliferation.
    Hernando H, Islam AB, Rodríguez-Ubreva J, Forné I, Ciudad L, Imhof A, Shannon-Lowe C, Ballestar E
    Epstein-Barr virus (EBV) infects and transforms human primary B cells inducing indefinite proliferation. To investigate the potential participation of chromatin mechanisms during the EBV-mediated transformation of resting B cells we performed an analysis of global changes in histone modifications. We observed a rema...

    Genome-wide shRNA screening to identify factors mediating Gata6 repression in mouse embryonic stem cells.
    Cooper S, Brockdorff N
    The use of whole-genome pooled shRNA libraries in loss-of-function screening in tissue culture models provides an effective means to identify novel factors acting in pathways of interest. Embryonic stem cells (ESCs) offer a unique opportunity to study processes involved in stem cell pluripotency and differentiation....

    The histone methyltransferase ASH1 orchestrates fibrogenic gene transcription during myofibroblast transdifferentiation.
    Perugorria MJ, Wilson CL, Zeybel M, Walsh M, Amin S, Robinson S, White SA, Burt AD, Oakley F, Tsukamoto H, Mann DA, Mann J
    BACKGROUND AND AIMS: Transdifferentiation of hepatic stellate cells (HSCs) to a myofibroblast-like phenotype is a pivotal event that drives liver fibrosis. HSC transdifferentiation requires coordinated global changes in gene expression. Here we have investigated epigenetic regulators that orchestrate HSC transdiffer...

    RYBP-PRC1 Complexes Mediate H2A Ubiquitylation at Polycomb Target Sites Independently of PRC2 and H3K27me3.
    Tavares L, Dimitrova E, Oxley D, Webster J, Poot R, Demmers J, Bezstarosti K, Taylor S, Ura H, Koide H, Wutz A, Vidal M, Elderkin S, Brockdorff N
    Polycomb-repressive complex 1 (PRC1) has a central role in the regulation of heritable gene silencing during differentiation and development. PRC1 recruitment is generally attributed to interaction of the chromodomain of the core protein Polycomb with trimethyl histone H3K27 (H3K27me3), catalyzed by a second complex...

    Glutathione-S-transferase pi 1(GSTP1) gene silencing in prostate cancer cells is reversed by the histone deacetylase inhibitor depsipeptide.
    Hauptstock V, Kuriakose S, Schmidt D, Düster R, Müller SC, von Ruecker A, Ellinger J
    Gene silencing by epigenetic mechanisms is frequent in prostate cancer (PCA). The link between DNA hypermethylation and histone modifications is not completely understood. We chose the GSTP1 gene which is silenced by hypermethylation to analyze the effect of the histone deacetylase inhibitor depsipeptide on DNA meth...

    Epigenetic silencing mediated through activated PI3K/AKT signaling in breast cancer.
    Zuo T, Liu TM, Lan X, Weng YI, Shen R, Gu F, Huang YW, Liyanarachchi S, Deatherage DE, Hsu PY, Taslim C, Ramaswamy B, Shapiro CL, Lin HJ, Cheng AS, Jin VX, Huang TH
    Trimethylation of histone 3 lysine 27 (H3K27me3) is a critical epigenetic mark for the maintenance of gene silencing. Additional accumulation of DNA methylation in target loci is thought to cooperatively support this epigenetic silencing during tumorigenesis. However, molecular mechanisms underlying the complex inte...

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