Elisa kit for il 6

Dulbecco's modified Eagle's medium (DMEM) was purchased from Flow Laboratories (Irvine, UK); glutamine and antibiotics were from Hyclone (Cramlington, UK). Insulin was from Sigma (St Louis, MO, USA). PCA was purchased from Sigma-Aldrich (St. Louis, MO, USA). Electrophoresis reagents were from Bio-Rad (Hercules, CA, USA). Anti-IRS-1 (catalog number sc-7200), anti-GLUT4 (catalog number sc-53566), anti-p65 NF-κB (catalog number sc-8008), IL-1β (catalog number sc-52865) and horseradish peroxidase-conjugate anti-goat (catalog number sc-7020), and anti-rabbit (catalog number sc-2357) antibodies were from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Horseradish peroxidase-conjugate anti-mouse (catalog number 172-1011) antibody was from Bio-Rad (Kidlington, UK). Anti-Akt (catalog number 2920S), anti-phospho Akt Ser 473 (catalog number 4060S), anti-IRβ (catalog number 81764), anti-phospho p65 NF-κB Ser536 (catalog number 3033S) antibodies were from Cell Signaling Technology (Beverly, MA, USA). Antibody against PTP1B (catalog number Ab2009) was purchased from Abcam (Cambridge, United Kingdom). Anti-phospho Tyr 608 IRS-1 (catalog number 09-432) and anti-β-actin (catalog number MABT826) antibodies were from Merck-millipore (Temecula, CA, USA). ELISA Kit for IL-6 was from Biolegend (San Diego, CA, USA) and Tyrosine Phosphatase Assay System from Promega (Madison, WI, USA).

Intestinal tissues of mice infected with S. Typhimurium were washed and transferred into protease inhibitor (Complete, Roche) containing NP40 buffer at indicated time-points. The tissue was disrupted mechanically using bead-beating four times for 1 min with 1mm zirconia/silica beads. Sample were stored at À80 C until analysis. Concentrations cytokines were measured using commercial ELISA kit for IL-6 (BioLegend) and multiplex assays for all other cytokines (ProcartaPlex, eBioscience and Legendplex kit, Biolegend). ELISA and multiplex kits were used according to manufacturer's instructions.