Diagenode

H2BK12ac polyclonal antibody - Classic

Antibody ChIP-seq grade icon
Catalog Number
Format
Price
C15410212
50 μg
$305.00
  Bulk order
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Polyclonal antibody raised in rabbit against the region of histone H2B containing the acetylated lysine 12 (H2BK12ac), using a KLH-conjugated synthetic peptide.

LotA2260P
Concentration0.82 µg/µl
Species reactivityHuman
TypePolyclonal
PurityAffinity purified
HostRabbit
PrecautionsThis product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications Suggested dilution References
ChIP * 0.5 - 1 μg per ChIP Fig 1, 2
ELISA 1:1,000 Fig 3
Dot Blotting 1:5,000 Fig 4
Western Blotting 1:1,000 Fig 5
Immunofluorescence 1:500 Fig 6
* Please note that the optimal antibody amount per IP should be determined by the end-user. We recommend testing 0.5-5 μg per IP.
  • Validation Data

    ChIP

    Figure 1. ChIP results obtained with the Diagenode antibody directed against H2BK12ac
    ChIP assays were performed using human HeLa cells, the Diagenode antibody against H2BK12ac (Cat. No. C15410212) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (cat. No. C01010051), using sheared chromatin from 1.5 million 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 a region approximately 1 kb upstream of the GAPDH and ACTB promoters, used as positive controls, and for the coding region of the inactive MYOD1 gene 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).

    ChIP-seq figure A

    ChIP-seq figure B

    ChIP-seq figure C

    ChIP-seq figure D

    Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against H2BK12ac
    ChIP was performed on sheared chromatin from 1.5 million HeLaS3 cells using 0.5 μg of the Diagenode antibody against H2BK12ac (Cat. No. C15410212) as described above. The IP’d DNA was subsequently analysed on an Illumina HiSeq. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 51 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the enrichment along the complete sequence and a 1 Mb region of the X-chromosome (fig 2A and B) and in genomic regions of chromosome 7, surrounding the ACTB gene, and of chromosome 12, surrounding the GAPDH gene (fig 2C and D). The position of the amplicon used for ChIP-qPCR is indicated by an arrow.

    ELISA

    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 H2BK12ac (Cat. No. C15410212) 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 1:38,200.

    Dot Blot analysis

    Figure 4. Cross reactivity tests using the Diagenode antibody directed against H2BK12ac
    To test the cross reactivity of the Diagenode antibody against H2BK12ac (Cat. No. C15410212), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H2B. 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.

    Western blot

    Figure 5. Western blot analysis using the Diagenode antibody directed against H2BK12ac
    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 H2BK12ac (Cat. No. C15410212). The antibody was diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The marker (in kDa) is shown on the left.

    Immunoflurescence

    Figure 6. Immunofluorescence using the Diagenode antibody directed against H2BK12ac
    HeLa cells were stained with the Diagenode antibody against H2BK12ac (cat. C15410212) 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 H2BK12ac 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
    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
    ChIP-seq (ab)
    Read more
    ChIP-qPCR (ab)
    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
  •  Documents
    Datasheet H2BK12ac C15410212 DATASHEET
    Datasheet description
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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...
    Download
  •  Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: H2BK12ac polyclonal antibody - Classic (Diagenode Cat# C15410212 Lot# A2260P). Click here to copy to clipboard.

    Using our products in your publication? Let us know!

    Myc Regulates Chromatin Decompaction and Nuclear Architecture during B Cell Activation
    Kieffer-Kwon K.R. et al.
    50 years ago, Vincent Allfrey and colleagues discovered that lymphocyte activation triggers massive acetylation of chromatin. However, the molecular mechanisms driving epigenetic accessibility are still unknown. We here show that stimulated lymphocytes decondense chromatin by three differentially regulated steps. Fi...

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