Sandwich elisa

Fasting blood sugar (FBS) was carried out using Cobas C 311, Roche/Hitachi USA and HbA1c levels were measured by Roche Cobas e-50, Switzerland. The analysis for antioxidants was carried out through sandwich ELISA (Bioassay Technology Laboratory, Shanghai, China) for SOD (Cat No. E0918Hu), GPX (Cat No. E3696Hu), NO (Cat No. E1510Hu), Vit C (Cat No. E1538Hu) while TAC (Cat No. E-BC-K225) was measured by Colorimetric Assay Kit (FRAP Method) by Elabscience USA. Uric acid was measured by TBHBA DIALAB Production, Austria.

After a 12 hour overnight fast, 5 cm³ blood was obtained for serum analyses. After centrifugation at 3000 rpm for 5 min, metabolic parameters were analysed immediately, but serum FFAs and LBP were analysed after supplying in −80°C.
FBS was measured by the enzymatic colorimetric method using glucose oxidase. Serum TG concentration was measured by commercially available enzymatic reagents with glycerol phosphate oxidase. Serum HDL-C was measured after precipitation of the apolipoprotein B-containing lipoproteins with phosphotungistic acid. Assays were performed using Pars Azmoon kits (Pars Azmoon, Tehran, Iran) and a Selectra 2 auto-analyzer (Vital Scientific, Spankeren, the Netherlands). Interassay and intra-assay coefficient of variation (CV) was <5% for all assays. Serum samples for both LBP and FFAs assays were stored at −80°C until analysis. Both serum LBP and FFAs levels were determined by a sandwich ELISA (Bioassay Technology Laboratory, Shanghai Korean Biotech, Shanghai City, China) according to the manufacturer’s instructions. The intra-assay and interassay CVs were <8% and <10%, respectively.