Epiquik hdac activity assay kit

The effect of SFN on HDAC activity in HeLa cells was determined using the EpiQuik HDAC activity assay kit (Epigentek, USA) according to the manufacturer's instructions. Briefly, HeLa cells were treated with SFN and TSA for 3 days and harvested and nuclear extracts were then prepared for the same using EpiQuik nuclear extraction kit (Epigentek, USA) following the manufacturer's instructions.

The EpiQuik HDAC Activity Assay Kit (Catalog Number P-4002, Epigentek, United States) was used for evaluating the effect of chrysin on the HDAC activity. The acetylated histone substrate–coated assay plate was prepared by adding 50 μL of the biotinylated HDAC substrate diluted in wash buffer to all wells. The assay plate was incubated at room temperature for 45 min and washed with wash buffer. HDAC assay buffer was dispensed to the wells in chrysin (5, 10, and 15 µM) treated and untreated nuclear extract placed. The plate was kept at 37°C for an hour followed by incubation with capture and detection antibody. Developing solution was used to develop the signal, stop solution stopped the reaction, and OD was read at 450 nm. The percentage inhibition of the HDAC activity against chrysin treatment was calculated by comparing with DMSO control. The experiment was repeated three times, and mean ± SD was used to plot a graph. One-way ANOVA was used to check the statistical significance at the p-value ≤ 0.05.