Rabbit anti il 1β pab

Western blotting was performed as described previously [30 (link)]. The following primary antibodies were used: mouse anti-β-actin mAb (1:5000; Sigma-Aldrich, A5441); rabbit anti-IL-1β pAb (1:1000; Abcam, ab9722); rabbit anti-TNF-α pAb (1:1000; Abcam, ab9739); rabbit anti-GAPDH pAb (1:5000; Proteintech, 10494-1-AP); rabbit anti-alpha-synuclein pAb (1:2000; Cell Signaling, 2628); rabbit anti-TNFR2 pAb (1:1000; Beyotime, AF8199); rabbit anti-HA tag pAb (1: 1000; Abcam, ab9110); mouse anti-FLAG tag mAb (1: 5000; Sigma-Aldrich, F1804); The membrane was washed and incubated for 2 h at room temperature with the corresponding secondary antibodies: (a) HRP-conjugated goat anti-rabbit IgG (1:10,000; Jackson ImmunoResearch Laboratories, 115-035-003); (b) HRP-conjugated goat anti-mouse IgG (1:10,000; Jackson ImmunoResearch Laboratories, 115-035-003). Peroxidase activity was detected with SuperSignal WestPico chemiluminescent substrate (Pierce Biotechnology) and visualized and digitized with ImageQuant (LAS-4000, Fujifilm, Japan). Optical densities of bands were analyzed by using ImageReader software (Fujifilm, Japan). Protein levels, quantified by computer analysis as the ratio between each immunoreactive band and the levels of β-actin or GAPDH, were expressed as a percentage of vehicle-treated control.

Western blotting was performed as described previously [32 (link)]. The following primary antibodies were used: (a) rabbit anti-IL-1β pAb (1:1000; Abcam, ab9722), (b) mouse anti-β-actin mAb (1:5000; Sigma-Aldrich, A5441), (c) rabbit anti-phosphor-SMAD2 pAb (1:1000; Invitrogen #51-1300), (d) rabbit anti-SMAD2 pAb (1:250; Cell signaling #3108), (e) rabbit anti-TNF-α pAb (1:100; Abcam, ab9739), and (f) mouse anti-Olig1 mAb (1:1000; Millipore, mab5540). The membrane was washed and incubated for 1 h at room temperature with the corresponding secondary antibodies: (a) HRP-conjugated goat anti-rabbit IgG (1:10,000; Jackson ImmunoResearch Laboratories, 115-035-003) and (b) HRP-conjugated goat anti-mouse IgG (1:10,000; Jackson ImmunoResearch Laboratories, 115-035-003). Peroxidase activity was detected with SuperSignal WestPico chemiluminescent substrate (Pierce Biotechnology) and visualized and digitized with ImageQuant (LAS-4000, Fujifilm, Japan). Optical densities of bands were analyzed by using ImageJ software (NIH, USA). Protein levels, quantified by computer analysis as the ratio between each immunoreactive band and the levels of β-actin, were expressed as a percentage of vehicle-treated control.