Plasma Collection and Cytokine Quantification

Venous blood samples, after fasting for 12h, were obtained with anticoagulant Ethylenediaminetetraacetic acid (EDTA), centrifuged at 3000 rpm for 15 min; further, plasma and buffy-coat were separated, divided into aliquots and stored at -80°C until thawed for assays, according to Flauzino et al.¹² (link) The IL-10 and TGF-β1 plasma levels were determined using microspheres multiplex immunofluorimetric assay (Procarta Plex High Sensitivity Assay by Thermo Fisher Scientific, Vienna, Austria) for the Luminex platform (MAGPIX™, Luminex Corp., Austin, TX, USA), that was performed according to the manufacturer's recommendations, as described in Stadlober et al.¹⁶ (link)