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Figure 1. ChIP results obtained with the Diagenode monoclonal antibody directed against H4K20me2 ChIP assays were performed using HeLa cells, the monoclonal antibody against H4K20me2 (Cat. No. C15200220) and optimized PCR primer sets for qPCR. ChIP was performed with the Auto Histone ChIP-seq kit (Cat. No. C01010022) on the IP-Star automated system, using sheared chromatin from 1 million cells. Respectively 1 and 5 μg of antibody was used per ChIP experiment. IgG (1 μg/IP) was used as negative IP control. QPCR was performed with primers for the promoters of the GAPDH and EIF4A2 genes, used as negative controls, and for the Sat2 and SatA satellite repeats, used as positive 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. Cross reactivity tests using the Diagenode monoclonal antibody directed against H4K20me2 To test the cross reactivity of the Diagenode monoclonal antibody against H4K20me2 (Cat. No. C15200220), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H4K20. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. Figure 2 shows a high specificity of the antibody for the modification of interest.
Figure 3. Western blot analysis using the Diagenode monoclonal directed antibody against H4K20me2 Histone extracts (15 μg) from HeLa cells were analysed by Western blot using the Diagenode monoclonal antibody against H4K20me2 (Cat. No. C15200220) 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.
Figure4. Immunofluorescence using the Diagenode monoclonal antibody directed against H4K20me2 HeLa cells were stained with the Diagenode monoclonal antibody against H4K20me2 (Cat. No. C15200220) and with DAPI. Cells were fixed with 4% formaldehyde for 10’ and blocked with PBS/TX-100 containing 1% BSA. The cells were immunofluorescently labeled with the H4K20me2 antibody (middle) diluted 1:1.000 in blocking solution followed by an anti-mouse antibody conjugated to Alexa594. The left panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.
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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Immunofluorescence using the Diagenode monoclonal antibody directed against CRISPR/Cas9
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