Acetyl-Histone H3 (Lys27) Antibody #4353

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-It is highly critical that the chromatin is of appropriate size and concentration.	-For optimal ChIP results, use 20 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. -It is important to keep the tissue cold to avoid protein degradation. -Use fresh formaldehyde that is not past the manufacturer's expiration date.	-Once in solution, store 1M DTT at -20°C.

Upstream tips
-It is highly critical that the chromatin is of appropriate size and concentration.
Protocol tips
-For optimal ChIP results, use 20 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. -It is important to keep the tissue cold to avoid protein degradation. -Use fresh formaldehyde that is not past the manufacturer's expiration date.
Downstream tips
-Once in solution, store 1M DTT at -20°C.

Publication protocol

HCT116 cells were treated with 30 μM JMF3086 for 24 h, and ChIP assays using control Rabbit IgG (sc-2027; Santa Cruz) or anti-H3K27-ac antibodies (#4353; Cell Signaling) were performed as previously described (Chou et al., 2011). ChIP-on-chip assays were performed in the Microarray and Gene Expression Analysis Core Facility of the National Yang-Ming University VGH Genome Research Center in Taiwan using the SurePrint G3 Human Promoter 1 × 1 M Kit (Agilent Technologies) after DNA was further purified through phenol-chloroform-isoamyl alcohol extraction and ethanol precipitation. Genes with two-fold downregulation or upregulation of H3K27-Ac in log-ratio after JMF3086 treatment were regarded significant.

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Manufacturer protocol

Download the product protocol from Cell Signaling Technology for Acetyl-Histone H3 (Lys27) Antibody #4353 below.

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