β actin antibody 4970

Liquiritin (CAS: 551-15-5, molecular formula is shown in Figure 1(a)) and fluoxetine hydrochloride (CAS: 56296-78-7), purity ≥98%, were purchased from Selleck China (Shanghai, China). PC12 cell was purchased from Procell Life Science&Technology Co., Ltd. DMSO (D4540) was got from Sigma-Aldrich (Shanghai) Trading Co., Ltd. Superoxide dismutase (SOD) detection kit (A001-1-2), glutathione peroxidase (GSH-Px) detection kit (A005-1-2), and malondialdehyde (MDA) detection kit (A003-1-2) were all purchased from Nanjing Jiancheng Institute of Biological Engineering (Nanjing, China). BCA protein concentration detection kit (P0012S) and RIPA Lysis Buffer (P0013B) were purchased from Beyotime Biotechnology Co., Ltd. (Shanghai, China). Interleukin-1 beta (IL-1β) detection kit (SEKM-0002), interleukin-18 (IL-18) detection kit (SEKM-0019), NLRP3, and TLR4 antibody (K004108P, K003881P) were all purchased from Beijing Solarbio Technology Co., Ltd. (Beijing, China). Caspase-1, pro-caspase-1, IL-1β, NEK7, and ASC (ab138483, ab238972, ab283818, ab133514, ab175449) antibodies were obtained from Abcam (Shanghai) Trading Co., Ltd. (Shanghai, China). β-Actin antibody (4970) and goat anti-rabbit IgG (HRP polymer) (98164) were purchased from Cell Signaling Technology (Danvers, USA).

Cells were treated as indicated and then lysed by RIPA (Beyotime Biotechnology, Shanghai, China). Protein concentration was quantified with the Bicinchoninic Acid Kit (Sigma-Aldrich, Shanghai, China) according to the manufacturer’s instructions. Equal amounts of protein (50 μg) were loaded for SDS-PAGE electrophoresis, transferred to a polyvinylidene fluoride microporous membrane (Millipore, Billerica, MA). The signals were probed with primary antibodies and amplified by the secondary antibodies. The primary antibodies included ARG1 (DF6657, Affinity Biosciences, Cincinnati, OH, USA), iNOS (18985–1-AP, Proteintech, Rosemont, IL, USA), CXCL1(AF5403, Affinity Biosciences, Cincinnati, OH, USA), CXCR2 (20634–1-AP, Proteintech, Rosemont, IL, USA), β-actin antibody (4970, Cell Signaling Technology, Danvers, MA, USA), MMP2 (A6247, ABclonal Technology Cambridge, Boston, USA), MMP9 (10375–2-AP, Proteintech, Rosemont, IL, USA). Finally, the bands were imaged through the ECL Advance reagent (Tanon Science & Technology, Shanghai, China).