Il 1β elisa kit

Lico.A, a pale yellow powder with 99% purity, was purchased from Pufei De Biotech (Chengdu, China). The Trizol reagent, Poly-L-lysine (PLL), Apomorphine, Griess reagent, [³H]DA in Krebs–Ringer buffer, LPS from Escherichia coli, O55:B55, and Dimethyl sulfoxide (DMSO) were obtained from Sigma-Aldrich (St. Louis, MO, USA). The 0.25% trypsin and Penicillin–streptomycin (PS) were purchased from Invitrogen (Carlsbad, CA, USA). The Dulbecco’s modified Eagle’s medium (DMEM), minimum essential medium (MEM), fetal bovine serum (FBS) and horse serum (HS) were purchased from Gibco (Grand Island, NY, USA). The PrimeScript^® 1st Strand cDNA Synthesis Kit was purchased from Takara Biotechnology (Dalian, China). The SYBR Green QuantiTect RT-PCR Kit was purchased from Roche (South San Francisco, CA, USA). The PGE2, TNF-α, IL-6 and IL-1β ELISA kits were purchased from R&D Systems (Abingdon, UK).

Tofacitinib (S5001, Selleck, Texas, USA), fludarabine (S1491, Selleck, Texas, USA), TUNEL reaction solution (ab66108, Abcam, Cambridge, UK), TTP488 (HY-50682, MedChem Express, Sollentuna, Sweden), LPS (L8880, Solarbio, Beijing, China), caspase-1 ELISA kits (KL15200, KALANG, Shanghai, China), IL-1β ELISA kits (RLB00, R&D Systems, Minneapolis, USA), IL-18 LISA kits (ab213909, Abcam, Cambridge, UK), Anti-NLRP1 antibody (ab3683, Abcam, Cambridge, UK), anti-NeuN antibody (ab177487, Abcam, Cambridge, UK), Anti-caspase-1 antibody (BM4291, Boster, Wuhan, China), Anti-IL-1β antibody (ab9722, Abcam, Cambridge, UK), Anti-IL-18 antibody (PB0058, Boster, Wuhan, China), anti-β-actin antibody (Boster, BA2305, Wuhan, China), followed by secondary antibodies conjugated to horseradish peroxidase anti-rabbit IgG (H+L) (AS014, ABclonal, Wuhan, China), and anti-mouse IgG (H+L) (AS003, ABclonal, Wuhan, China) were used in this study.

The levels of IL-1β protein in cell-free supernatants were determined using IL-1β ELISA kits (Cat#: DY401-05, R&D systems) according to the manufacturer’s instructions.

Cell culture media were collected, and the secreted interleukin-10 (IL-10) and interleukin-1β (IL-1β) protein concentrations were measured by ELISAs. The culture medium was centrifuged at 4°C for 15 min at 12,000 g to collect the supernatant, which was then assayed and quantified with IL-10 and IL-1β ELISA kits (R&D Systems, USA) according to the manufacturer’s instructions.

The anti-Lyn, anti-Src, anti-α-tubulin antibodies and horseradish peroxidase conjugated secondary antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). The anti-mouse TNF-α, IL-6, and IL-1β ELISA kits were obtained from R&D Systems (Minneapolis, MN). Anti-phospho-tyrosine (4G10) antibody was from Upstate (Temecula, CA), and anti-pLyn (Tyr 396) antibody was purchased from Abcam (Cambridge, MA). Anti-pSrc, Src, pERK, pJNK, p-p38, JNK, p38 and ERK2 antibodies were purchased from Cell Signaling Technology (Danvers, MA). LPS (Lipopolysaccharides from E.Coli 026:B6) was obtained from Sigma-Aldrich Corp. (St. Louis, MO). The LDH cytotoxicity assay kit was from Promega Corporation (Madison, WI). Human Aβ1–42 was purchased from rPeptide (Bogart, GA).

Baicalin was obtained from Nanjing Zelang Medical Technology. LPS, dimethyl sulfoxide (DMSO), 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), indomethacin (INDO), dexamethasone (DEX), aminoguanidine (AG, iNOS inhibitor), NS-398 (COX-2 inhibitor), and pyrrolidine dithiocarbamate (PDTC, IκBα, and NF-κB P65 inhibitor) were purchased from Sigma-Aldrich. NO and iNOS assay kits were purchased from Nanjing Jiancheng Bioengineering Institute. Real-time PCR kit was purchased from Shanghai Gene Pharma. PGE2 and IL-1β ELISA kits were purchased from R&D Systems. MAPK family antibody sampler kit, phosphorylated MAPK family antibody sampler kit, and primary antibodies against β-actin, COX-2, IRAK1, IRAK4, MyD88, and NF-κB P65 were purchased from Cell Signaling Technology. Interleukin-1 receptor-associated kinase 1/4 inhibitor (IRAK1/4 inhibitor), TLR4, P-IκBα, and IκBα antibodies were purchased from Santa Cruz Biotechnology. MTS510 (TLR4 inhibitor) was purchased from eBioscience. SB203580 (P38 MAPK inhibitor), SP600125 (JNK inhibitor), ST2825 (MyD88 inhibitor), and U0126 (ERK inhibitor) were purchased from the Beyotime Institute of Biotechnology. RPMI 1640 and fetal bovine serum (FBS) were purchased from Gibco.