Epiquik global histone h4 acetylation assay kit

The effects of TNFα alone or in combination with HCA on H3 and H4 histone acetylation were evaluated with the EpiQuik™ Global Histone H3 Acetylation Assay Kit and EpiQuik™ Global Histone H4 Acetylation Assay Kit (Epigentek, Farmingdale, NY, USA). Following the manufacturer’s instructions, histones were extracted from HH that had been treated for 24 h with 5 ng/mL TNFα alone or combined with 500 μM HCA. The histones were spotted onto the wells, and then an antibody specific for acetylated histone H3 or H4 was added. After washing the wells, HRP-conjugated secondary antibody was added, followed by the detection reagent, and absorbance was measured with GloMax® Discover Microplate Reader at 450 nm. Percent acetylation was calculated as OD (treated sample − blank)/OD (untreated control − blank) × 100%.

Histone H3 and H4 acetylation were quantified using the EpiQuik™ Global Histone H3 Acetylation Assay Kit (Epigentek, Farmingdale, NY, USA) and the EpiQuik™ Global Histone H4 Acetylation Assay Kit (Epigentek, Farmingdale, NY, USA) according to the manufacturer’s instructions. First, histones were isolated from freshly isolated PBMCs using the reagents of the respective kit as well as trichloroacetic acid (TCA) and acetone. Histones were then quantified using the Pierce™ Coomassie Plus Bradford Assay (Thermo Scientific™, Waltham, MA, USA). Histone extracts were stored at −80 °C until assayed. Subsequently, 1 ug of protein was immobilized on a 96-well plate and acetylated H3/H4 was detected with capture and detection antibodies. The plate was read at 450 nm using the FlexA-200 microplate reader (Allsheng, Hangzhou, China), and OD was displayed directly.