Diagenode

H4K12ac polyclonal antibody

Catalog Number
Format
Price
C15410331-50
50 μg
$380.00
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Polyclonal antibody raised in rabbit against the region of histone H4 containing the acetylated lysines 12 (H4K12ac), using a KLH-conjugated synthetic peptide. 

LotA2439P
Concentration0.6 μg/μl
Species reactivityHuman, mouse, wide range expected
TypePolyclonal
PurityAffinity purified
HostRabbit
Storage ConditionsStore at -20°C; for long storage, store at -80°C. Avoid multiple freeze-thaw cycles.
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP/ChIP-seq * 0.5-1 μg/ChIP Fig 1, 2
ELISA 1:1,000 Fig 3
Dot Blotting 1:1,000 Fig 4
Western Blotting 1:500 Fig 5
IF 1:200 Fig 6

* 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
    H4K12ac Antibody ChIP Grade

    Figure 1. ChIP results obtained with the Diagenode antibody directed against H4K12ac
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H4K12ac (Cat. No. C15410331) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (Cat. No. C01010055) on sheared chromatin from 1,000,000 cells. A titration of the antibody consisting of 0.5, 1, 2 and 5 μg per ChIP experiment was analysed. IgG (1 μg/IP) was used as negative IP control. QPCR was performed with primers for promoter of the active GAPDH and EIF4A2 genes, used as positive controls, and for the inactive MYOD1 gene and the Sat2 satellite repeat region 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).

    A. H4K12ac Antibody ChIP-seq Grade

    B. H4K12ac Antibody for ChIP-seq

    C. H4K12ac Antibody for ChIP-seq assay

    D. H4K12ac Antibody validated in ChIP-seq

    Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against H4K12ac
    ChIP was performed with 0.5 μg of the Diagenode antibody against H4K12ac (Cat. No. C15410331) on sheared chromatin from 1,000,000 HeLa cells using the “iDeal ChIP-seq” kit as described above. The IP’d DNA was subsequently analysed on an Illumina HiSeq 2000. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the peak distribution along the complete sequence and a 2 Mb region of the human X chromosome (figure 2A and B) and in two genomic regions surrounding the GAPDH and EIF4A2 positive control genes (figure 2C and D).

    H4K12ac Antibody ELISA validation

    Figure 3. 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 H4K12ac (Cat. No. C15410331) 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 3), the titer of the antibody was estimated to be 1:14,300.

    H4K12ac Antibody Dot Blot validation

    Figure 4. Cross reactivity tests using the Diagenode antibody directed against H4K12ac
    To test the cross reactivity of the Diagenode antibody against H4K12ac (Cat. No. C15410331), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H4. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:1,000. Figure 4 shows the antibody is specific for the K12 acetylation with some slight cross reaction with K8ac.

    H4K12ac Antibody validated in Western Blot

    Figure 5. Western blot analysis using the Diagenode antibody directed against H4K12ac
    Western blot was performed on whole cell (25 μg, lane 1) and histone extracts (15 μg, lane 2) from HeLa cells, and on 1 μg of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the Diagenode antibody against H4K12ac (Cat. No. C15410331). The antibody was diluted 1:500 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.

    H4K12ac Antibody validated in Immunofluorescence

    Figure 6. Immunofluorescence using the Diagenode antibody directed against H4K12ac
    HeLa cells were stained with the Diagenode antibody against H4K12ac (Cat. No. C15410331) 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 labeled with the H4K12ac antibody (left) diluted 1:200 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.

  • Target Description

    Histones are the main constituents of the protein part of chromosomes of eukaryotic cells. They are rich in the amino acids arginine and lysine and have been greatly conserved during evolution. Histones pack the DNA into tight masses of chromatin. Two core histones of each class H2A, H2B, H3 and H4 assemble and are wrapped by 146 base pairs of DNA to form one octameric nucleosome. Histone tails undergo numerous post-translational modifications, which either directly or indirectly alter chromatin structure to facilitate transcriptional activation or repression or other nuclear processes. In addition to the genetic code, combinations of the different histone modifications reveal the so-called “histone code”. Histone methylation and demethylation is dynamically regulated by respectively histone methyl transferases and histone demethylases. Acetylation of histone H4 is associated with active genes.

  •  実験手法
    ChIP-seq (ab)
    Read more
    ChIP-qPCR (ab)
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    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
  •  資料
    H4K12ac polyclonal antibody DATASHEET
    Polyclonal antibody raised in rabbit against the region of histone H4 containing the acetylated l...
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    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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  •  Safety sheets
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  •  出版物

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H4K12ac polyclonal antibody (Diagenode Cat# C15410331-50 Lot# A2439P). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

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