ChIPseq results obtained with ab195478 directed against Histone H3 (acetyl K56).

ChIP was performed on sheared chromatin from 1.5 million HeLaS3 cells using 5 µg of ab195478. The IP’d DNA was subsequently 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 shows the enrichment along the complete sequence and a 1 Mb region of the X-chromosome (fig A and B) and in genomic regions of chromosome 12 and 3, surrounding the GAPDH and EIF4A2 positive control genes.

ChIP results obtained with ab195478 directed against Histone H3 (acetyl K56).

ChIP assays were performed using human HeLa cells, ab195478 and optimized PCR primer sets for qPCR. ChIP was performed 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 GAPDH promoter and for the EIF4A2 promoter, used as positive controls, and for the coding region of the inactive MYOD1 gene and the Sat2 satellite repeat, used as negative controls. Figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

Dot Blot analysis was performed to test the cross reactivity of ab195478 against Histone H3 (acetyl K56) with peptides containing other histone modifications and the unmodified H3K56. 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 shows a high specificity of the antibody for the modification of interest.