Diagenode

H3K27me1 polyclonal antibody - Classic

RFX5-polyclonal-antibody-diagenode
Catalog Number
Format
Price
C15410045
(pAb-045-050)
50 µg/82 µl
$295.00
  Bulk order

Polyclonal antibody raised in rabbit against histone H3 containing the monomethylated lysine 27 (H3K27me1), using a KLH-conjugated synthetic peptide.

LotA932-00234P
Concentration1.93 µg/µl
Species reactivityHuman, Arabidopsis
TypePolyclonal
PurityAffinity purified
HostRabbit
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP * 1 μg/ChIP Fig 1
ELISA 1:500 Fig 2
Dot Blotting 1:20,000 Fig 3
Western Blotting 1:1,000 Fig 4
Immunofluorescence 1:1,000 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 H3K27me1
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H3K27me1 (Cat. No. pAb-045-050) and optimized PCR primer sets for qPCR. ChIP was performed with the “LowCell# ChIP” kit (Cat. No. kch-maglow-016), using sheared chromatin from 100,000 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 and the coding region of the active gene GAPDH used as a negative and a positive control target, respectively. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

    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 H3K27me1 (Cat. No. pAb-045-050). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution (Figure 1), the titer of the purified antibody was estimated to be 1:32,900.

    Dot Blot

    Figure 3. Cross reactivity tests using the Diagenode antibody directed against H3K27me1
    A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K27me1 (Cat. No. pAb-045-050) with peptides containing other modifications and unmodified sequences of histone H3 and H4. One hundred to 0.2 pmol of the peptide containing the respective histone modification 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 directed against H3K27me1
    Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode antibody against H3K27me1 (Cat. No. pAb-045-050) 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 A

    Immunofluorescence figure B

    Immunofluorescence figure C

    Immunofluorescence figure D

    Immunofluorescence figure E

    Figure 5. Immunofluorescence using the Diagenode antibody directed against H3K27me1
    Human osteosarcoma (U2OS) cells were stained with the Diagenode antibody against H3K27me1 (Cat. No. pAb-045-050) and with DAPI. Cells were fixed with 4% formaldehyde for 20’ and blocked with PBS/TX-100 containing 5% normal goat serum. Figure 5A: cells were immunofluorescently labeled with the H3K27me1 antibody (left) diluted 1:1,000 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa568 or with DAPI (right), which specifically labels DNA. Figure 5B, C, D and E: staining of the cells with the H3K27me1 antibody after incubation of the antibody with 2 ng/μl blocking peptide containing the unmodified and the mono-, di- and trimethylated H3K27, respectively.

  • Applications
    ELISA
    Enzyme-linked immunosorbent assay. Read more
    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
  • Documents
    Datasheet H3K27me1 pAb-045-050 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: H3K27me1 polyclonal antibody - Classic (Diagenode Cat# C15410045 Lot# A932-00234P). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Parental epigenetic asymmetry of PRC2-mediated histone modifications in the Arabidopsis endosperm
    Moreno-Romero J et al.
    Parental genomes in the endosperm are marked by differential DNA methylation and are therefore epigenetically distinct. This epigenetic asymmetry is established in the gametes and maintained after fertilization by unknown mechanisms. In this manuscript, we have addressed the key question whether parentally inherited...

    A novel microscopy-based high-throughput screening method to identify proteins that regulate global histone modification levels.
    Baas R, Lelieveld D, van Teeffelen H, Lijnzaad P, Castelijns B, van Schaik FM, Vermeulen M, Egan DA, Timmers HT, de Graaf P
    Posttranslational modifications of histones play an important role in the regulation of gene expression and chromatin structure in eukaryotes. The balance between chromatin factors depositing (writers) and removing (erasers) histone marks regulates the steady-state levels of chromatin modifications. Here we describe...

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