Circulating NLRP3 and Cytokine Levels

Circulating levels of NLRP3 was estimated using enzyme-linked immunosorbent assays (ELISA), catalog number CSB-E15885h, Cusabio, Houston, TX, USA. According to the manufacturer’s protocol, the minimum detectable dose for this assay was <0.039 ng/mL of human NLRP3 with a CV% of <8% and <10% for intra- and inter-assay precision, as previously described [8 (link)]. Circulating levels of IL-1β, IL-18 and IL-37 were assessed using the Flex MAP-3D System (Luminex Corporation, Austin, TX, USA), which utilized human cytokine magnetic bead panels (IL-18 and IL-1β: HCYTA-60K human cytokine, chemokine growth factor panel A—immunology multiplex assay) (IL-37: HCYP4MAG-64K human cytokine chemokine magnetic bead panel IV). The intra- and inter-assay % CVs for IL-18, IL-1β and IL- 37 were <15 and <20 and <10 and <15, respectively. Circulating levels of IL-1α and IL-33 were estimated using ELISA assays (Bio Vendor, R & D systems, Brno, Czech Republic) Cat No. RAF045R and RAF064R, respectively. The intra-assay % CVs for both were <5.4% and 4.7%, respectively, while the inter-assay % CVs for both were <10% and 6.9%, respectively. The controls and standards implemented in all the biochemical assays were regularly reviewed by the quality assurance team of CBCD in KSU.