| Assay Method Information | |
| | Fluorometric Assay |
| Description: | Compounds were tested for their ability to inhibit histone deacetylase 8 using an in vitro deacetylation assay. In a detailed procedure, 8 μl of HDAC8 enzyme solution (0.125 μg/μl) was transferred to assay plates (CORNING 3676). 1 μl of half-log diluted compounds were added into wells and incubated for 15 min at rt. After that, 8 μl of substrate solution was transferred to the wells and incubated for 30 min at rt. After deacetylation of the substrate by incubation with HDAC8 enzyme, subsequent exposure to a developing reagent produced a fluorophore that was directly proportional to the level of deacetylation. So 4 μl of developer regent (Cayman 10006394) was added into the wells in the assay plates and incubated for 15 min. The fluorescence signal was measured by a FlexStation3 plate reader (excitation wave length 340-360 nm; emission wave length 440-465 nm). |
| Affinity data for this assay | |
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