M9410

Histone methyl-lysine peptides, listed in Table 2, were serially diluted in high-bind, hydrophobic 96-well plates (Sigma, #M9410) and allowed to incubate 37 ℃, 100 rpm, overnight. The following day, plates were washed with 1X PBS twice, and blocked with 1X PBS containing 4% BSA for 2 h, 37 ℃. Following blocking, plates were washed twice with 1X PBS containing 0.15% Tween-20. All antibodies were diluted 1:10,000 in 1X PBS (containing 4% BSA and 0.15% v/v Tween-20), added to plates and allowed to incubate at 37 ℃ for 2 h. Peptide competition reactions were pre-incubated 2 h at room temperature with the respective antibody prior to addition to the ELISA plates. Following incubation with primary antibody incubation, the plates were washed 3X with 1X PBS containing 0.15% v/v Tween-20 and incubated with secondary antibody (1:10,000) for 2 h at 37 °C. Following incubation in secondary antibody, the plates were washed 3X with 1X PBS containing 0.15% v/v Tween-20. The plates were then incubated with buffer containing dibasic sodium phosphate (20 mM), citric acid (10 mM) and fresh hydrogen peroxide (final concentration: 0.0012%), and incubated for 30 min at room temperature in the dark. The reaction was quenched with 1 M H₂SO₄ and imaged on a spectrometer at 492 nm. All data are represented in Additional file 1: Fig. S6.