Cytokines and MPO-DNA quantification
Blood was collected into EDTAK2 anticoagulant (10% v/v). After centrifugation of whole blood at 1000G for 5 minutes at 4℃, plasma was collected, and the cell pellet was removed. NET-associated MPO was captured on coated 96-well plates from the mouse MPO ELISA kit (Abcam, Cambridge, UK, Roche, Basel, Switzerland). Cytokines: Interleukin (IL)-6, IL-10, Interferon-γ (IFN-γ), Tumor Necrosis Factor-alpha (TNF-α), Macrophage Inflammatory Protein 2 (MIP-2) and Monocyte Chemoattractant Protein-1(MCP-1) were measured using Luminex ® xMAP according to the manufacturer’s instructions. A standard curve was used to calculate the concentrations of MPO-DNA and cytokines.