Mouse il 17 quantikine elisa kit

Blood samples were obtained by cardiac puncture and collected in heparinized tubes [36 (link),60 (link)]. BAL samples were obtained according to the technique developed in the Laboratory Immunobiotechnology of CERELA-CONICET (San Miguel de Tucuman, Argentina) [38 (link),61 (link)]. The trachea was exposed and intubated with a catheter, and then two sequential bronchoalveolar lavages were performed on each mouse by injecting sterile PBS. The recovered fluid was centrifuged for 10 min at 900× g and the fluid frozen at −70 °C for subsequent cytokine determinations. Tumor necrosis factor alpha (TNF-α), interferon gamma (IFN-γ), interleukin 10 (IL-10), IL-1β, IL-6, IL-17, KC, and monocyte chemoattractant protein 1 (MCP-1) concentrations were determined in serum and BAL samples using commercial ELISA kits. IFN-γ (Mouse IFN-gamma Quantikine ELISA Kit, sensitivity: 2 pg/mL), IL-10 (Mouse IL-10 Quantikine ELISA Kit, sensitivity: 5.2 pg/mL), IL-1β (Mouse IL-1β DuoSet ELISA, sensitivity: 1.5 pg/mL), IL-6 (Mouse IL-6 Quantikine ELISA Kit, sensitivity: 1.8 pg/mL), and IL-17 (Mouse IL-17 Quantikine ELISA Kit, sensitivity: 4.7 pg/mL) from R&D Systems (USA). TNF-α (Mouse TNF alpha ELISA Kit, sensitivity: 9.1 pg/mL), MCP-1 (Mouse MCP1 ELISA Kit, sensitivity: 0.487 pg/mL), and KC (Mouse KC ELISA Kit, sensitivity: 1.95 pg/mL) kits were obtained from Abcam (Cambridge, UK).

Serum of NOD/Ltj mice was collected and separated from heart blood for cytokine analysis. The following kits were used in this study: mouse IFNγ Quantikine ELISA kit (MIF00, R&D Systems, US) and mouse IL-17 Quantikine ELISA kit (M1700, R&D Systems, US). The procedure was performed according to the manufacturer's instructions.

The 1.5-cm small intestine tissues were cut from the distal ileum of each mice, opened longitudinally, washed in ice-cold PBS immediately, and weighed. Then, the tissues were lysed with T-PER™ Tissue Protein Extraction Reagent (Thermo Scientific™, USA) supplemented with protease inhibitors (Yeasen, China), followed by centrifugation at 1,500 g for 20 min at 4°C. The supernatants from the histiocyte lysates were collected and used immediately for subsequent experiments or stored at −80°C. IL-17A and IL-6 concentrations were measured using Mouse IL-17 Quantikine ELISA Kit and Mouse IL-6 Quantikine ELISA Kit (R&D Systems, USA). Cytokine concentrations in the tissues were expressed as amount per gram of tissue. Cytokine concentrations in the serum were expressed as amount per mL of serum.