Western blot was performed using histone extracts from HeLa cells (HeLa HE, 20 ug) and H3K27me2/3 monoclonal antibody (Cat # MAB4947) at dilution 1 : 1,000 in TBS-Tween + 5% skimmed milk. The location of the protein of interest is indicated with an arrow.
Dot Blot is performed to determine the cross-reactivity of H3K27me2/3 monoclonal antibody (Cat # MAB4947) with other histone modifications. Other histone modifications include mono-methylation of lysine 27 and mono-, di- and trimethylation of two other lysines of H3 (K9 and K36). To determine the cross-reactivity, 0.2 to 100 pmol of peptide containing the respective histone modifications are spotted on a membrane. The antibody is used at a dilution of 1 : 300. The antibody recognizes di- and trimethylation of lysine 27 on histone H3.