| Assay Method Information | |
| | In vitro HDACs Inhibition Fluorescence Assay |
| Description: | In brief, 10 μL of enzyme solution (HeLa nuclear extracts, HDAC1, HDAC4, HDAC6, HDAC8, HDAC11) was mixed with various concentrations of target compounds (50 μL), SAHA, and PXD101, using 100% and none HDACs groups as control group. Afterincubation at 37 °C for 10 min, 40 μL of fluorogenic substrate (Boc-Lys(Ac)-AMC for HeLa nuclear extracts; Ac-Leu- GlyLyS(Ac)-AMC for HDAC1, HDAC6, and HDAC11; Ac-Leu-GlyLyS(Tfa)-AMC for HDAC4 and HDAC8) was added, and then, the mixture was incubated at 37 °C for 30 min. The mixture was stopped by addition of 100 μL of developer containing trypsin and TSA afterward. Over the next incubation at 37 °C for 20 min, fluorescence intensity was measured using a microplate reader at excitation and emission wavelengths of 390 and 460 nm, respectively. |
| Affinity data for this assay | |
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