H3K14ac Antibody
Polyclonal antibody raised in rabbit against the region of histone H3 containing the acetylated lysine 14 (H3K14ac), using a KLH-conjugated synthetic peptide.
| Lot | A2283P |
|---|---|
| Concentration | 0.9 µg/µl |
| Species reactivity | Human |
| Type | Polyclonal |
| Purity | Affinity purified |
| Host | Rabbit |
| Precautions | This product is for research use only. Not for use in diagnostic or therapeutic procedures. |
| Applications | Suggested dilution * | References |
|---|---|---|
| ChIP * | 2 μg/ChIP | Fig 1 |
| ELISA | 1:1,000 - 1:5,000 | Fig 3 |
| Dot Blotting | 1:10,000 | Fig 3 |
| Western Blotting | 1:1,000 | Fig 4 |
* 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
Figure 1. ChIP results obtained with the Diagenode antibody directed against H3K14ac
ChIP assays were performed using human HeLa cells, the Diagenode antibody against H3K14ac (cat. No. C15410310) and optimized PCR primer sets for qPCR. ChIP was performed with the “iDeal ChIP-seq” kit (cat. No. C01010055), 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 ACTB and GAPDH 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).
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 H3K14ac (cat. No. C15410310) 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 2), the titer of the antibody was estimated to be 1:73,200.
Figure 3. Cross reactivity tests using the Diagenode antibody directed against H3K14ac
To test the cross reactivity of the Diagenode antibody against H3K14ac (cat. No. C15410310), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H3K14. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:10,000. Figure 3 shows a high specificity of the antibody for the modification of interest.
Figure 4. Western blot analysis using the Diagenode antibody directed against H3K14ac
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 H3K14ac (cat. No. C15410310). The antibody was 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.
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 H3 is associated with active genes.
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-qPCR (ab) Read more |
Documents
| Datasheet H3K14ac C15410310 DATASHEET Datasheet description | Download |
Safety sheets
| SDS C15410310 H3K14ac Antibody US en | Download |
| SDS C15410310 H3K14ac Antibody GB en | Download |
| SDS C15410310 H3K14ac Antibody FR fr | Download |
| SDS C15410310 H3K14ac Antibody BE nl | Download |
| SDS C15410310 H3K14ac Antibody ES es | Download |
| SDS C15410310 H3K14ac Antibody BE fr | Download |
| SDS C15410310 H3K14ac Antibody JP ja | Download |
| SDS C15410310 H3K14ac Antibody DE de | Download |
Publications
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CRISPR-based precise methylation of specific FUT8 promoter regions allows isolation of CHO cells with a fine-tuned glycoprofile |