Luminex-based Multiplexed Cytokine Assay

Cytokine levels were determined using a multiplexed bead immunoassay and measured with a Luminex MAGPIX instrument (Luminex, Austin, TX, USA) as previously detailed [22 (link), 23 (link)]. In brief, TNF‐α and CXCL10 were measured using magnetic bead assay (Mouse XL Cytokine Luminex Performance Premixed Kit; Bio‐Techne, Minneapolis, MN, USA) following manufacturer's instructions. Each assay was analyzed on the Luminex MAGPIX instrument to measure inflammatory concentrations followed by a 5‐parameter logistic curve‐fitting method from a standard curve of each respective analyte. Plasma samples were normalized by equivalent volume of commercially supplied dilution buffer and examined in duplicate per each assay run. Concentration was calculated by the statlia Immunoassay Analysis software (Brendan Bioanalytics, Carlsbad, CA, USA).