Milliplex map human cytokine chemokine growth factor panel

Serum samples frozen and stored at -30°C without other thawing were tested for simultaneous quantification of sCD40L, EGF, eotaxin, FGF-2, FLT-3L, fractalkine, G-CSF, GM-CSF, GRO-α, IFN-α2, IFN-γ, IL-1α, IL-1β, IL-1RA, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12 (p40), IL-12 (p70), IL-13, IL-15, IL-17A, IL-17E/IL-25, IL-17F, IL-18, IL-22, IL-27, IP-10, MCP-1, MCP-3, M-CSF, MDC, MIG, MIP-1α, MIP-1β, PDGF-AA, TGF-α, TNF-α, TNF-β and VEGF-A with MILLIPLEX MAP Human Cytokine/Chemokine/Growth Factor Panel (Merck-Millipore). All assays were performed according to the manufacturer’s protocol for serum samples, utilizing recommended sample dilutions and standard curve concentrations (Merck-Millipore). Acquisitions were performed using a MAGPIX instrument operated with xPONENT software version 4.2 (Luminex Corp.).

Plasma cytokine IL-6, IL-10 and TNF-α levels were assessed using the multiplex bead-based immunoassay Luminex system using the Milliplex^® MAP Human Cytokine/Chemokine/Growth Factor panel (EMD Millipore, Billerica, MA, USA) according to the manufacturers’ protocol. Analytes included IL-6, IL-10, and TNF-α. Twenty-five microliters of neat plasma sample was used in the wells, and the plate was sealed and incubated with agitation on a plate shaker overnight (16–18 h) at 2–8 °C. Data were analyzed using Luminex xPonent 3.1 software. The average CVs of quality control were 6.30 ± 4.26% for IL-6, 13.57 ± 15.68% for IL-10 and 8.03 ± 2.71% for TNF-α.