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Assay Method Information |
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| | Scintillation Proximity Assay (SPA) |
| Description: | Experiments were performed in triplicate at room temperature with 1 h incubation of 10 μl reaction mixture in buffer of 20 mM Tris-HCl, pH 8.0, 5 mM DTT, and 0.01%Triton X-100 containing 30 nM of SUV420H1 or 250 nM of SUV420H2, 2 μM of [3H]SAM (Cat. # NET155V250UC; PerkinElmer; http://www.perkinelmer.com) and 8 μM of cold SAM and 3 μM of peptide H4K20Me1. To stop the enzymatic reactions, 10 μl of 7.5 M guanidine hydrochloride was added, followed by 180 μl of buffer (20 mM Tris, pH 8.0), mixed and then transferred to a 96-well FlashPlate (Cat. # SMP103; PerkinElmer). After mixing, the reaction mixtures in Flashplate were incubated for 2 h and the counts per minute (CPM) were measured using TopCount plate reader (PerkinElmer). |
| Affinity data for this assay | |
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