Quantikine hs elisa human il 6 immunoassay

Before, immediately after and 90 minutes after completion of the ultra-marathon, blood samples were taken for determination of the plasma concentrations of IL-6, IL-10, IL-18, IL-1β and tumour necrosis factor alpha (TNF-α). The blood samples were collected in tubes containing EDTA. All samples were centrifuged within 15 minutes at 3000 rpm at 4°C, and stored at -70°C until further processed. High-sensitivity ELISA kits provided by R&D Systems (Minneapolis, MN, USA) were used for the pre-race blood samples for IL-6, IL-10 and TNF-α (Quantikine HS ELISA Human IL-6 Immunoassay, Quantikine HS ELISA Human IL-10 Immunoassay and Quantikine HS ELISA Human TNF-α Immunoassay, respectively) because these cytokines exist at very low levels in peripheral blood. Quantitative sandwich ELISA kits were used for the post-race and recovery blood samples for IL-6, IL-10, IL-18, IL-1β and TNF-α (R&D Systems Inc.; Minneapolis, MN, USA). Plasma concentration of IL-18 was determined using a Human IL-18 Instant ELISA kit (eBioscience, Bender MedSystems GmbH, Austria). All samples and provided standards were analysed in duplicate. Haematocrit was measured in duplicate. Whole blood samples (∼9 µl) were transferred to heparinized microcapillary tubes and analysed by an automated system following microcentrifugation. Haematocrit was used to calculate percent changes in plasma volume.

We used an immunoturbidimetric assay with the ADVIA 1800 Chemistry System to measure serum levels of high-sensitivity CRP; the inter- and intraassay coefficients of variation (CVs) were 10.8% and 5.6%, respectively. Plasma levels of TNF receptor 1 (TNFR1) were measured using an immunoassay kit (Human sTNF RI/TNFRSF1A Immunoassay, R&D Systems Inc.); the inter- and intraassay CVs were 7.4%–11.0% and 9.6%, respectively. Plasma levels of IL-6 were measured using a high-sensitivity ELISA (Quantikine HS ELISA Human IL-6 Immunoassay, R&D Systems) per the manufacturer’s instructions; the inter- and intraassay CVs were 10.3%–15.3% and 16.6%, respectively.
An overall inflammatory marker score was created by summing the z-scores of the above-mentioned log-transformed inflammatory biomarkers: z-score (log high-sensitivity CRP) + z-score (logTNFR1) + z-score (logIL-6) [11 (link),26 (link)].