H3K79me3 Antibody
Polyclonal antibody raised in rabbit against histone H3 containing the trimethylated lysine 79 (H3K79me3), using a KLH-conjugated synthetic peptide.
| Lot | A2014P |
|---|---|
| Concentration | 0.53 µg/µl |
| Species reactivity | Human, mouse, yeast: positive. Other species: not tested. |
| Type | Polyclonal |
| Purity | Affinity purified polyclonal antibody |
| Host | Rabbit |
| Storage Conditions | Store at -20°C; for long storage, store at -80°C. Avoid multiple freeze-thaw cycles. |
| Storage Buffer | PBS containing 0.05% azide and 0.05% ProClin 300. |
| Precautions | This product is for research use only. Not for use in diagnostic or therapeutic procedures. |
| Applications | Suggested dilution | References |
|---|---|---|
| ChIP/ChIP-seq * | 1-2 μg/ChIP | Fig 1, 2 |
| ELISA | 1:500 – 1/1,000 | Fig 3 |
| Dot Blotting | 1:20,000 | Fig 4 |
| Western Blotting | 1:1,00 | Fig 5 |
* 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
Figure 1. ChIP results obtained with the Diagenode antibody directed against H3K79me3
ChIP was performed with the Diagenode antibody against H3K79me3 (cat. No. C15410068) on sheared chromatin from 1 million HeLaS3 cells using the “Auto Histone ChIP-seq” kit and the SX-8G IP-Star automated system. A titration of the antibody consisting of 1, 2, 5 and 10 µg per ChIP experiment was analysed. IgG (5 µg/IP) was used as negative IP control. Quantitative PCR was performed with primers for the GAPDH promoter and for exon 2 of the inactive myoglobin gene. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis). These results are in accordance with the observation that H3K79me3 shows a preference for active promoters.
Figure 2. ChIP-seq results obtained with the Diagenode antibody directed against H3K79me3
ChIP was performed on 100,000 K562 cells using 1 µg of the Diagenode antibody against H3K79me3 (cat. No. C15410068). The IP'd DNA was analysed on an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. Figure 2 shows the peak distribution along the complete sequence and a 3 Mb region of human chromosome 1 (figure 2A and B), in a 2 genomic regions surrounding the GAPDH and EIF2S3 positive control genes (figure 2C and D).
Figure 3. 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 H3K79me3 (cat. No. C15410068) 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 1:8,500.
Figure 4. Cross reactivity tests using the Diagenode antibody directed against H3K79me3
A Dot Blot analysis was performed to test the cross reactivity of the Diagenode antibody against H3K79me3 (cat. No.C15410068) with peptides containing other histone modifications and the unmodified H3K79. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:100,000. Figure 4 shows a high specificity of the antibody for the modification of interest.
Figure 5. Western blot analysis using the Diagenode antibody directed against H3K79me3
Histone extracts of HeLa cells (15 µg) were analysed by Western blot using the Diagenode antibody against H3K79me3 (cat. No.C15410068), diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The molecular weight marker is shown on the right; the location of the protein of interest is indicated on the left.
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. Trimethylation of histone H3 on K79 was shown to be more present at active promotors than at silent promotors.
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 |
Documents
| Datasheet H3K79me3 C15410068 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... | Download |
| Epigenetic Antibodies Brochure BROCHURE More than in any other immuoprecipitation assays, quality antibodies are critical tools in many e... | Download |
Safety sheets
| SDS C15410068 H3K79me3 Antibody GB en | Download |
| SDS C15410068 H3K79me3 Antibody US en | Download |
| SDS C15410068 H3K79me3 Antibody BE nl | Download |
| SDS C15410068 H3K79me3 Antibody FR fr | Download |
| SDS C15410068 H3K79me3 Antibody JP ja | Download |
| SDS C15410068 H3K79me3 Antibody DE de | Download |
| SDS C15410068 H3K79me3 Antibody BE fr | Download |
| SDS C15410068 H3K79me3 Antibody ES es | Download |
Publications
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Distinct silencer states generate epigenetic states of heterochromatin. |
DOT1L O-GlcNAcylation promotes its protein stability andMLL-fusion leukemia cell proliferation. |
The histone methyltransferase DOT1L is required for proper DNA damage response, DNA repair, and modulates chemotherapy responsiveness. |
Tri-methylation of H3K79 is decreased in TGF-β1-induced epithelial-to-mesenchymal transition in lung cancer |
DOT1L Activity Promotes Proliferation and Protects Cortical Neural Stem Cells from Activation of ATF4-DDIT3-Mediated ER Stress In Vitro |
MLL-Rearranged Acute Lymphoblastic Leukemias Activate BCL-2 through H3K79 Methylation and Are Sensitive to the BCL-2-Specific Antagonist ABT-199 |
Degree of recruitment of DOT1L to MLL-AF9 defines level of H3K79 Di- and tri-methylation on target genes and transformation potential. |
Anticheckpoint pathways at telomeres in yeast |
Promoter-exon relationship of H3 lysine 9, 27, 36 and 79 methylation on pluripotency-associated genes. |
Chromatin states of core pluripotency-associated genes in pluripotent, multipotent and differentiated cells. |