Legend max human mrp8 14 elisa kit

Venous blood samples were drawn from all patients in the emergency department before the initiation of any treatment as standard-of-care, and neutrophils, lymphocytes (Alinity hq, Abbott, USA), and serum glucose (Architect i2000SR, Abbott, USA) were measured by standard laboratory techniques. After receiving or not treatment, new blood samples were taken within the following 24 h and C-reactive protein (CRP) was measured with an autoanalyzer (Architect i2000SR, Abbott, USA). These second samples were centrifuged at 1200×g for 15 min, 4 °C within 2 h of collection, and stored at -80°C for further analysis. Citrated plasma samples were thawed on ice and thoroughly vortexed before measuring calprotectin levels (LEGEND MAX Human MRP8/14 ELISA Kit, BioLegend, USA) with an automated ELISA analyzer TRITURUS (Grifols, Spain). The inter- and intra-assay variability averages were 7.4% and 2.8%, respectively. All experiments were performed in a blinded manner following the manufacturer’s instructions.

The pro-inflammatory S100A8/A9 complex and cytokine TNF-α in the cell culturing medium were measured by Legend MAX Human MRP8/14 ELISA kit (BioLegend, San Diego, CA, USA) and Human TNF-α ELISA Max Deluxe sets (BioLegend), respectively, according to the manufacturer’s instructions.

Calprotectin levels in feces were detected by ELISA according to the manufacturer’s protocol (Legend max human MRP8/14 ELISA kit, Biolegend). In brief, 50 mg frozen feces were dissolved in 1 mL PBS for 30 min and vortexed for 5 min. The homogenized samples were centrifuged with 3,000 × g for 5 min at 4°C, followed by 10,000 × g for 10 min at 4°C. The supernatant was removed and stored at −80°C until used. Calprotectin concentration was determined at an optical density at 450 nm, and the results were determined from the standard curve, including a total sample dilution of 1:2,500. Serum samples for ASCA analysis were collected from an independent cohort of patients with HD and healthy controls following the same screening criteria. The serum samples were collected and stored at −80°C until used. Levels of ASCA in serum (n = 20 for AE, n = 26 for CE, and n = 45 for healthy control) were quantified by enzyme immunoassay following the manufacturer’s protocol (ASCA IgG ELISA kit, Abnova). The level of ASCA (U/mL) was measured at 450-nm wavelength, and a sample with an ASCA level higher than 20 U/mL was defined as positive.