| Assay Method Information | |
| | AlphaScreen Assay |
| Description: | Binding assays were carried out as 20-μl reactions in 384-well white ProxiPlates (Perkin-Elmer Life Sciences) as described [Hong et al., Proc. Natl. Acad. Sci. U.S.A., 104:18439-18444]. His-tagged enzyme (500 nM) was incubated with biotinylated H3K27Me3 peptide (Biotin-KAPRKQLATKAARKMe3SAPATGG, variable concentration) for 15 min at room temperature in buffer containing 50 mM HEPES (pH 7.5), 0.01% Tween 20, 0.1% BSA. AlphaScreen streptavidin-conjugated donor and nickel chelate-conjugated acceptor beads were added to the wells at a final concentration of 10 μg/ml and incubated for a further 1 h in the dark at 22 °C. The plates were analyzed using an Envision (PerkinElmer Life Sciences) plate reader. |
| Affinity data for this assay | |
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