Rat il 4 elisa kit

Levels of cytokines in the liver were determined by specific ELISA kit assay (Rat TNFα ELISA kit, # 88-7340-88, eBioscience; rat IL-1β ELISA kit, DY501; rat IL-6 ELISA kit, DY506; rat IL-4 ELISA kit, DY504; rat IL-10 ELISA kit, DY522, all from R&D Systems) [32] (link). Cytokine measurement was done in whole cell extracts from the livers isolated by previously published protocol [33] (link). In brief, 100 mg of liver was homogenized in 10 vol. (w/v) of PBS buffer (137 mM NaCl, 2.7 mM KCl, 10 mM Na 2 HPO 4 , 1.8 mM KH 2 PO 4 , 0.5% Triton X-100 and protease inhibitors), then sonicated 3 × 5 s/30 s pause (30 Hz, pulse 0.5). Homogenates were centrifuged at 12,000×g, 10 min, 4 °C, and the resulting supernatants were aliquoted and stored at -70 °C. The level of cytokines was measured in 96-well plates according to the manufacturer's instructions. Absorbance was read at 450 and 540 nm on the Multiskan Spectrum (Thermo Electron Corporation, Finland). Results were expressed as pg/ mg of protein.