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Figure 1. ChIP results obtained with the Diagenode monoclonal antibody directed against H4K16ac ChIP assays were performed using HeLa cells, the monoclonal antibody against H4K16ac (Cat. No. C15200219) 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 the coding region of the EIF2S3 gene, used as a positive control, and for the Sat2 satellite repeat, used as a negative control. 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. Cross reactivity tests using the Diagenode monoclonal antibody directed against H4K16ac To test the cross reactivity of the Diagenode monoclonal antibody against H4K16ac (Cat. No. C15200219), a Dot Blot analysis was performed with peptides containing other acetylations and unmodified sequences of histone H4. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:10,000. Figure 2 shows a high specificity of the antibody for the modification of interest.
Figure 3. Western blot analysis using the Diagenode monoclonal antibody directed against H4K16ac Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode monoclonal antibody against H4K16ac (Cat. No. C15200219) 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.
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.
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