Image forming system

The expression levels of proteins were performed using western blot. Tissue extracts were prepared using RIPA buffer. After electrophoresis, proteins were transferred onto PVDF membranes. Then the membrane was blocked for 1 h at room temperature with 5% milk solution, and incubated with primary antibodies at 4°C overnight. The following antibodies were used in this study: NF-κB (6956S, Cell Signaling Technology), pNF-κB (3033, Cell Signaling Technology), Mx1 (ab79609, Abcam), IFN-α (ab230993, Abcam), HSP70 (ADI-SPA-810-F, Enzo Life Sciences), I-FABP (sc-16063, Santa Cruz Biotechnology), Caspase-3 (9661S, Cell Signaling Technology), Villin (sc-7672, Santa Cruz Biotechnology), AQP4 (ab46182, Abcam), AQP3 (ab125219, Abcam), Claudin (RF217968, Invitrogen), and Occludin (TC259714, Invitrogen), β-actin (PA1-46296, Invitrogen). Subsequently, appropriate HRP-conjugated secondary antibodies were used followed by incubation at room temperature for 1 h. Blots were visualized using a chemiluminescence kit (Amersham Biosciences, Uppsala, Sweden) and image forming system (Alpha Innotech, New York, NY, USA).

The expression levels of proteins were performed using western blotting as described by Hou et al. [64 (link)]. The primary antibodies: HSP70, Caspase-3, Bax, Villin, and Occludin (rabbit, 1:1000; Cell Signaling Technology, Inc., Danvers, MA, USA), β-actin (mouse 1:2000; Sigma-Aldrich Inc., St. Louis, MI, USA). The secondary antibody: Anti-rabbit (mouse, 1:2000; Zhongshan Golden Bridge Biological Technology Co., Ltd., Beijing, China). Blots were carried out by utilizing a chemiluminescence kit (Amersham Biosciences, Uppsala, Sweden) and image forming system (Alpha Innotech, New York, NY, USA).