Quantiglo elisa human tnfα immunoassay

To quantify the secretion of pro- (IL-6, MCP-1, chemerin, and TNFα) and anti-inflammatory (adiponectin) adipokines, cells were treated with bioactive compounds for 48 h and conditioned medium was collected for adipokine quantification by ELISA according to the manufacturer’s instructions. The samples were diluted 5-60 fold for IL-6 detection (Quantikine ELISA Human IL-6 Immunoassay; R&D Systems, Minneapolis, MN, Intra-assay precision: CV <5 %, Inter-assay precision: CV <4 %), 3-10 fold for MCP-1 (Quantikine ELISA Human CCL3/MCP-1 Immunoassay; R&D Systems, Intra- and Inter-assay precision: CV <6 %) and 2-10 fold for chemerin (Quantikine ELISA Human Chemerin Immunoassay; R&D Systems, Intra-assay precision: CV ≤3 %, Inter-assay precision: CV <6 %) to match the standard curve. Adiponectin (Mercodia Adiponectin ELISA; Mercodia AB, Uppsala, Sweden, Intra-assay precision: CV <5 %, Inter-assay precision: CV <8 %) and TNFα (Quantiglo ELISA Human TNFα Immunoassay; R&D Systems, Intra-assay precision: CV <6 %, Inter-assay precision: CV <9 %) secretion were measured in undiluted samples. Levels of TNFα were detectable in adipocyte cultures treated with AC from only two subjects.

A blood sample was obtained from each patient at Fukushima Medical University before discharge under a fasting state. The plasma B-type natriuretic peptide (BNP) levels were measured using a specific immunoradiometric assay (Shionoria BNP kit, Shionogi, Osaka, Japan). The troponin I was measured in EDTA anticoagulated plasma using a refined assay (Abbott-Architect; Abbott Laboratories, Abbott Park, Illinois, USA). Tumour necrosis-factor (TNF)-α was measured, based on the method of solid phase chemiluminescent ELISA using an immunoassay kit (Quanti Glo ELISA Human TNF-α Immunoassay, R&D Systems, Minneapolis, USA). Total serum adiponectin was based on the method of latex agglutination turbidimetric immunoassay, also using an immunoassay kit (Human Adiponectin Latex Immunoassay, LSI Medience, Tokyo, Japan).