Anti p tyr705 stat3

H7402 cells and HepG2 cells were plated at a density of 1 × 10⁶ cells in 10-cm dishes and cultured with 2 μM SAHA for 48 h. Chromatin immunoprecipitation (ChIP) was performed using the EZ ChIP kit (Millipore, Temecula, CA, USA) according to the manufacturer's recommendations. Briefly, cells were lysed and crosslinked with 1% formaldehyde. Lysates were sonicated to shear DNA to lengths between 200 and 1000 base pairs. An aliquot of the chromatin preparation was set aside and designated as the input fraction. Immunoprecipitations were carried out in ChIP Dilution Buffer with the following antibodies: IgG, anti-acetyl histone H3 (AcH3), anti-acetyl histone H4 (AcH4; Millipore), and anti-p-Tyr705-STAT3 (Cell Signaling Technology, Danvers, MA, USA). Immunoprecipitates were collected using protein G-sepharose beads, washed and eluted according to the manufacturer's protocol. Immunoprecipitated DNA was recovered by reversing crosslinking, and purified using the QIAGEN Purification Kit (QIAGEN, Hilden, Germany) and analysed by PCR. The sequences of the PCR primers used for MICA, miR-17-92 or MCM7 promoter analysis are listed in supplementary Table 3. Negative control primers (Millipore) amplified a non-related GAPDH promoter sequence.