Elecsys 1010 2010

In all patients, peripheral venous blood samples from the antecubital vein were collected between 7.00 and 8.00 a.m. after overnight fasting. Routine biochemical and hematological assessments were performed at the day of collection. Serum insulin was determined with electrochemiluminiscence immunoassay kits (Elecsys) on Roche Elecsys 1010/2010 and modular analytics E170 immunoassay analyzers (Roche Diagnostics GmbH, Germany); plasma glucose was measured by the glucose oxidase method on a Beckman autoanalyzer. Insulin resistance was estimated by the HOMA-IR using the following formula: fasting serum insulin (mU.l -1 ) x fasting plasma glucose (mmol.l -1 )/22.5 (Matthews et al. 1985) .
Serum leptin, soluble leptin receptor (sOB-R) and adiponectin were determined by the enzyme-linked immunosorbent assays (DRG, Germany; BioVendor, Germany). Leptin to sOB-R ratio was used to calculate the free-leptin index (FLI).

Venous blood samples were obtained from the antecubital area by phlebotomy between the hours of 08:00 and 09:00 following 10-12 h of fasting. After these samples were allowed to clot, they were centrifuged for 10 minutes at 3000 g. The sera were separated and stored at -70°C until the analysis.
Plasma glucose levels were measured by enzymatic colorimetric assay (GOD-PAP, Roche Diagnostics, Mannheim). Plasma zinc and copper levels were measured by atomic absorption spectrophotometry (ContrAA700, Analytik Jena AG, Germany) with an intra-assay CV of 2.4% and an inter-assay CV of 3.1%. Electrochemoluminescence immunoassay was used to measure serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol, prolactin and 17-hydroxy-progesterone (Roche Elecsys 1010/2010, Roche Diagnostics, Mannheim, Germany). The serum concentrations of dehidroepiandrosterone sulphate (DHEAS), thyroid stimulating hormone (TSH), total testosterone, free testosterone and insulin were measured by radioimmunoassay (DSL Diagnostic Systems Laboratories, USA). The intra-assay CVs were 5.3%, 3.8%, 7.8%, 6.2%, 5.0%, 9.6%, 10.0%, 9.6%, 9.7% and 8.2% whereas the inter-assay CVs were 1.8%, 1.5%, 10.0%, 5.7%, 4.1%, 9.3%, 7.8%, 6.6%, 6.2% and 7.4% for FSH, LH, estradiol, prolactin, 17-hydroxy-progesterone, DHEAS, TSH, total testosterone, free testosterone and insulin respectively.

Peripheral venous blood samples were collected between 6-7 a.m. following an overnight 12 h fast and polysomnography. Blood sample was taken from the antecubital vein and after immediate centrifugation, aliquots of plasma and serum were stored at -70 °C until analysis. Fasting cholesterol, triglycerides, high-density lipoprotein (HDL) cholesterol, apolipoprotein A-I (ApoA-I), and apolipoprotein B (ApoB) were measured by routine enzymatic methods. Low-density lipoprotein (LDL) cholesterol was derived using the Friedewald equation. Serum insulin was determined with electrochemiluminiscence immunoassay kits (Elecsys) on Roche Elecsys 1010/2010 and modular analytics E170 immunoassay analyzers (Roche Diagnostics GmbH, Mannheim, Germany); plasma glucose was measured by the glucose oxidase method on a Beckman autoanalyzer. Insulin resistance was estimated by the homeostasis model assessment (HOMA-IR) using the following formula: fasting serum insulin (mU.l -1 ) x fasting plasma glucose (mmol.l -1 )/22.5 (Gayoso-Diz et al. 2013) . LBP levels were assessed using a commercially available kit based on murine monoclonal antibodies specific for human LBP (Human LBP ELISA, Abnova, Taipei City, Taiwan).