Western blot analysis

Human DPCs cultured in α-MEM were exposed to dental monomers (1mM HEMA, 5mM MMA and 1mM TEGDMA) in the absence or presence of 10 mM NAC for 24 h (n = 3) and lysed in RIPA buffer. Protein contents of the lysed cells were measured with the BCA Protein Assay Kit. Extracted proteins were loaded on 10% sodium dodecyl sulfate polyacrylamide gels, transferred to polyvinylidene fluoride membranes (Bio-Rad), and blocked with 5% nonfat milk powder. Membranes were incubated overnight with the following primary rabbit anti-human antibodies (Cell Signaling Technology, Danvers, MA): anti-Bcl-2, anti-Bax, anti-p53, anti-cleaved caspase-3, and actin. Membranes were incubated with goat anti-rabbit secondary antibodies. Protein signals were visualized by using the ECL Western Blotting Detection System (GE Healthcare, Piscataway, NJ, USA). Protein expression levels were normalized to actin by using Image-Pro Plus 5.0 Software (Media Cybernetics Inc., Bethesda, MD, USA).