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H3K4ac Antibody - ChIP-seq Grade

Catalog Number
Format
Price
C15410322-10
10 μg
$95.00
  Bulk order
Other format



Polyclonal antibody raised in rabbit against histone H3 acetylated at lysine 4 (H3K4ac), using a KLH-conjugated synthetic peptide. 

LotA2505P
Concentration1.1 μg/μl
Species reactivityHuman
TypePolyclonal
PurityAffinity purified polyclonal antibody
HostRabbit
Storage ConditionsStore at -20°C; for long storage, store at -80°C. Avoid multiple freeze-thaw cycles.
Storage BufferPBS containing 0.05% azide and 0.05% ProClin 300.
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP/ChIP-seq* 2 µg/ChIP Fig 1, 2
ELISA 1:1,000 - 1:10,000 Fig 3
Dot Blotting 1:5,000 Fig 4
Western Blotting 1:1,000 Fig 5
Immunofluorescence 1:200 Fig 6

* Please note that the optimal antibody amount per IP should be determined by the end-user.

  • Validation data

    H3K4ac Antibody ChIP Grade

    Figure 1. ChIP results obtained with the Diagenode antibody directed against H3K4ac
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H3K4ac (Cat. No. C15410322) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (Cat. No. C01010051), using sheared chromatin from 4 million cells. A titration of the antibody consisting of 1, 2, 5 and 10 µg per ChIP experiment was analysed. IgG (1 μg/IP) was used as negative IP control. QPCR was performed with primers for the GAPDH and EIF4A2 promoters, used as positive controls, and for the HBB promoter and the Sat2 satellite repeat, 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).

    H3K4ac Antibody ChIP-seq Grade

    H3K4ac Antibody for ChIP-seq

    H3K4ac Antibody for ChIP-seq assay

    H3K4ac Antibody validated in ChIP-seq

    Figure 2. ChIP-seq results obtained with the Diagenode monoclonal antibody directed against TAL1
    ChIP was performed with 1 µg of the Diagenode antibody against H3K4ac (Cat. No. C15410322) on sheared chromatin from 4 million HeLa cells 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 1.5 mb region of the X chromosome (figure 2A and B) and in two regions surrounding the GAPDH and EIF4A2 positive control genes (figure 2C and D).

    H3K4ac 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 H3K4ac (Cat. No. C15410322) 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 purified antibody was estimated to be 1:197,500.

    H3K4ac Antibody Dot Blot validation

    Figure 4. Cross reactivity test using the Diagenode antibody directed against H3K4ac
    A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K4ac (Cat. No. C15410322) with peptides containing other histone H3 and H4 modifications and the unmodified H3K4 sequence. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:5,000. Figure 4 shows a high specificity of the antibody for the modification of interest.

    H3K4ac Antibody validated in Western Blot

    Figure 5. Western blot analysis using the Diagenode antibody directed against H3K4ac
    Whole cell (25 μg, lane 1) and histone extracts (15 µg, lane 2) from HeLa cells were analysed by Western blot using the Diagenode antibody against H3K4ac (Cat. No. C15410322), diluted 1:1,000 in TBSTween containing 5% BSA. The marker (in kDa) is shown on the left, the position of the protein of interest is indicated on the right.

    H3K4ac Antibody validated in Immunofluorescence

    Figure 6. Immunofluorescence using the Diagenode antibody directed against H3K4ac
    HeLa cells were stained with the Diagenode antibody against H3K4ac (Cat. No. C15410322) 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 H3K4ac 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.

  •  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
    ChIP-seq (ab)
    Read more
    ChIP-qPCR (ab)
    Read more
  •  Documents
    H3K4ac polyclonal antibody DATASHEET
    Dtasheet of the H3K4ac polyclonal antibody
    Download
  •  Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H3K4ac Antibody - ChIP-seq Grade (Diagenode Cat# C15410322-10 Lot# A2505P). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    SIRT1-dependent epigenetic regulation of H3 and H4 histone acetylation in human breast cancer
    Khaldoun Rifaï et al.
    Breast cancer is the most frequently diagnosed malignancy in women worldwide. It is well established that the complexity of carcinogenesis involves profound epigenetic deregulations that contribute to the tumorigenesis process. Deregulated H3 and H4 acetylated histone marks are amongst those alterations. Sirtuin-1 (...

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