Anti rabbit or mouse horseradish peroxidase antibody

Total protein extracted from astrocytes was separated by 12% SDS-PAGE. The separated proteins were transferred to a nitrocellulose (NC) membrane and incubated with antibodies against GFAP (Proteintech, Rosemont, USA), Shh (Abcam), Ptch (Sigma-Aldrich), Smo (Sigma-Aldrich), Gli-1 (Sigma-Aldrich), GRP78 (Proteintech), PERK, eIF2 (Proteintech), phospho-eIF2 (EnoGene Biotech Co., New York, USA), IRE1 (Signalway Antibody, Baltimore, USA), phospho-IRE1 (Boster, Pleasanton, USA), CHOP (Proteintech), and β-actin (Proteintech). The NC membrane was washed with TBS/T three times and then incubated with a 1:10,000 dilution of anti-rabbit or mouse horseradish peroxidase antibody (Sigma-Aldrich). The bands were detected by ECL reagents (EMD Millipore, Burlington, USA) and captured by a ChemiDoc Imaging System (Bio-Rad). ImageJ software analysis was employed to detect the optical density of the target proteins.

The proteins of astrocytes were separated by 12% SDS-PAGE. The separated proteins were transferred to nitrocellulose membrane and incubated with antibodies against Shh (Sigma-Aldrich), Ptch (Sigma-Aldrich), Smo (Sigma-Aldrich), Gli-1 (Sigma-Aldrich), IL-1β (RayBiotech, Norcross, USA), IL-6 (RayBiotech), NF-κB (Sigma-Aldrich) and β-actin (Sigma-Aldrich). The membrane was washed with TBS/T three times and then incubated with a 1:10,000 dilution of anti-rabbit or mouse horseradish peroxidase antibody (Sigma-Aldrich). The reactive bands were detected by ECL reagents (EMD Millipore, Billerica, USA) and captured by a UVP BioSpectrum 600 Imaging System (Analytik Jena US, Upland, USA). ImageJ software analysis was used to detect the image densitometry of target proteins.