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Monoclonal antibody raised in rabbit against the region of histone H3 containing the acetylated lysine 27 (H3K27ac), using a KLH-conjugated synthetic peptide.
Lot
006
Concentration
1 μg/μl
Species reactivity
Human, wide range expected.
Type
Monoclonal ChIP grade/ChIP-seq grade
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 50% glycerol, 1% BSA and 0.09% azide.
Precautions
This product is for research use only. Not for use in diagnostic or therapeutic procedures.
Applications
Suggested dilution
References
ChIP/ChIP-seq *
0.5 - 1 µg per IP
Fig 1
Dot Blotting
1:2,000
Fig 2
Western Blotting
1:1,000
Fig 3
Immunofluorescence
1:1,000
Fig 4
*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.
Figure 1. ChIP results obtained with the Diagenode monoclonal antibody directed against H3K27ac ChIP was performed on sheared chromatin from 500000 HeLa cells using 0.5 µg of the Diagenode antibody against H3K4me3 (cat. No. C15410003) as described above. The IP'd DNA was subsequently analysed on an Illumina NovaSeq. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the peak distribution along the complete sequence and a 2 Mb region of the human X-chromosome (figure 2A and B) and in two regions surrounding the GAPDH and EIF4A2 positive control genes, respectively (figure 2C and D).
A.
B.
C.
D.
Figure 2. ChIP-seq results obtained with the Diagenode monoclonal antibody directed against H3K27ac ChIP was performed on HeLa cells using 0.5 µg of the Diagenode antibody against H3K4me2 (cat. No. C15410035). The IP'd DNA was 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 peak distribution along the complete sequence and a 1.5 Mb region of the human X-chromosome (figure 2A and 2B) and in 2 chromosomal regions surrounding the ACTB and GAPDH positive control genes (figure 2C and D, respectively).
Figure 3. Cross reactivity tests using the Diagenode monoclonal antibody directed against H3K27ac To test the cross reactivity of the Diagenode antibody against H3K27ac (cat. No. C15210016), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H3K27. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:2,000. Figure 3 shows a high specificity of the antibody for the modification of interest.
Figure 4. Western blot analysis using the Diagenode monoclonal antibody directed against H3K27ac Western blot was performed on whole cell extracts (40 µg) from HeLa cells using the Diagenode antibody against H3K27ac (cat. No. C15210016). The antibody was diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is shown on the right, the marker (in kDa) is shown on the left.
Figure 5. Immunofluorescence using the Diagenode monoclonal antibody directed against H3K27ac HeLa cells were stained with the Diagenode antibody against H3K27ac (cat. No. C15210016, red) diluted 1:1,000. Actin was stained with fluorescein phalladoin (green).
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