Toll-like Receptor Protein Expression Analysis

The cells were washed with PBS and the lysates were prepared as previously described (16 (link)). Protein quantification was performed using a DC^TM protein assay kit (Bio-Rad) and run on SDS-PAGE gels. The primary antibodies consisted of anti-TLR1, anti-TLR2 (R&D Systems), anti-TLR4 (Novus Biologicals), anti-TLR10 (Santa Cruz Biotechnology, Santa Cruz, CA, USA), phospho-IκBα #9246 (Cell Signaling), NF-κBα p65 sc-372 (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and anti-β-actin antibodies. HRP-labeled goat anti-mouse IgG 1706516 (Bio-Rad) and HRP-labeled goat anti-rabbit IgG 1706515 (Bio-Rad) were used as secondary antibodies.