Specificity Analysis of Histone Modification-Specific Antibodies or Reading Domains on Histone Peptide Arrays.

Histone posttranslational modifications (PTMs) have a crucial role in chromatin regulation and dynamics. They are specifically bound by so-called reading domains, which mediate the biological effects of histone PTMs. On a similar note, antibodies are invaluable reagents in chromatin biology for the detection, characterization, and mapping of histone PTMs. Despite these central roles in chromatin research and biology, the specificity of many antibodies and reading domains has been insufficiently characterized and documented. Here we describe in detail the application of the MODified™ Histone Peptide Array for the investigation of the binding specificity of histone binding antibodies or domains. The array contains 384 histone tail peptides carrying 59 posttranslational modifications in different combinations which can be used to study the primary binding specificity, but at the same time also allow to determine the combinatorial effect of secondary marks on antibody or reading domain binding.