Glucose analyzer 2

All blood sampling was performed at the General Clinical Research Center and analyzed at VUMC central laboratories. After blood drawn was performed, samples were transported on ice and centrifuged at 3000 rpm for 15 minutes before kept frozen at −80 °C. Plasma fasting glucose concentrations were analyzed by using the glucose oxidase method (Glucose analyzer 2; Beckman Coulter, Brea, CA). Concentrations of serum albumin, prealbumin, bicarbonate and intact parathyroid hormone (iPTH) were measured using standard methods at VUMC central laboratories. Serum leptin levels were performed at Diabetes Research Training Center (DRTC) hormone laboratory by using certified methods. High sensitivity C-reactive protein (hs-CRP) concentrations were measured by high-sensitivity particle-enhanced turbidimetric UniCel DxI Immunoassay system (Beckman Coulter). Plasma IL-6 levels were determined using cytometric bead arrays (Becton Dickinson, San Jose, CA). Plasma protein thiol groups were analyzed according to the procedure which previously published by Ellman ¹⁰ et al and as modified by Hu ^{11 (link)} et al.

All blood samples were collected in similar fashion and technique to that described in Hung et al. 2010 (8 (link)). Glucose concentrations were measured by using the glucose oxidase method and insulin by double-antibody RIA, (Glucose Analyzer 2; Beckman Coulter, Brea, CA, and DA RIA; Millipore, St. Charles, MO, respectively). C-reactive protein levels were measured using the UniCel Dxi Immunoassay System (Beckman Coulter, Brea, CA). Adiponectin and leptin were measured using the MILLIPEX MAP Human Serum Adiponectin Panel A kit (Millipore, Billerica, MA). All of the other measurements were performed using standard, certified, routine laboratory methods.

Plasma leptin and insulin concentrations were measured with ELISA kits (R&D Systems and Crystal Chem Inc., respectively), and plasma glucose concentrations were determined using the Beckman glucose analyzer 2.

Plasma leptin and insulin concentrations were measured with ELISA kits (R&D Systems and Crystal Chem Inc., respectively) and plasma glucose concentrations were determined using the glucose oxidation method (Beckman glucose analyzer 2).