SCN− concentrations were quantified by ion-exchange chromatography26 (link),87 (link). Serum samples were mixed 1:1 (v/v) with acetonitrile, followed by centrifugation (5000g, 4 °C, 7 min) to remove precipitated proteins. The supernatant was subsequently filtered through 0.2 μm centrifuge filters (5000g, 4 °C, 3 min; Pall Nanosep MF, 500 μL capacity). Samples (25 μL) were then injected on to an IonPac AS16 column (4 × 250 mm) via an AG16 guard column (4 × 50 mm) fitted into a Dionex chromatography system consisting of an AS3500 autosampler, GP40 gradient pump, ASRS 300 anion suppressor (4 mm) in AutoSuppression recycle mode, and an ED40 electrochemical detector with DS3 detector stabilizer. Ions were eluted over 12 min with 50 mM potassium hydroxide in water with a flow rate of 1.5 mL min−1 and a suppressor current of 186 mA. Peak areas were quantified using Chromeleon Chromatography Studio software (version 7), against a standard curve generated using commercial NaSCN (0–200 μM).
ANP, BNP, CRP, and galectin-3 were quantified using commercial kits (Atrial Natriuretic Peptide EIA Kit, Brain Natriuretic Peptide EIA Kit, and rat C-Reactive Protein ELISA Kit, all from Sigma-Aldrich; and rat Galectin-3 ELISA from Ray Biotech) in accordance with the manufacturer’s instructions.
ANP, BNP, CRP, and galectin-3 were quantified using commercial kits (Atrial Natriuretic Peptide EIA Kit, Brain Natriuretic Peptide EIA Kit, and rat C-Reactive Protein ELISA Kit, all from Sigma-Aldrich; and rat Galectin-3 ELISA from Ray Biotech) in accordance with the manufacturer’s instructions.
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