Elecsys insulin assay

The recruited subjects gave information on their maternal age and pre-pregnancy weight. The weight and height of patients during the third trimester of pregnancy and the fetal weight were measured by standard methods, and both body again and pre-pregnancy body mass index (BMI) expressed as weight before pregnancy divided by height square were calculated.
Blood samples were drawn after a 12 h overnight fast. Serum total cholesterol, HDL-cholesterol, LDL cholesterol and triglycerides were determined by the total cholesterol CHOD-PAP and triglyceride GPO-PAP kits (Roche, Mannheim, Germany). Apoplipoprotein A1, apoplipoprotein B, and adiponetcin concentrations were measured by enzyme-linked immunosorbent assay method (AssayPro, St. Charles, USA). The glycated haemoglobin (HbA_1c) was measured by a latex enhanced turbidimetric immunoassay using specific monoclonal antibodies. Plasma insulin was quantified using Elecsys insulin assay (Roche). To assess insulin sensitivity, the quantitative insulin check index (QUICKI-IS) was calculated as follow: QUIKI = 1/[log(I0) + log (G0)], where I0 is the fasting plasma insulin (μU/ml) and G0 is the fasting blood glucose concentration (mg/dl) [38 ].

Hemogram will be performed by automatically counting blood cells from whole blood samples. Plasma glucose levels will be determined by the colorimetric glucose oxidase method. Blood will also be collected to assess the serum insulin levels and lipid profile. Serum insulinemia will be evaluated by an electrochemiluminescent immunoassay (Elecsys Insulin Assay, Roche Diagnostics GmbH, Mannheim, Germany) through an automatic analyzer (Cobas e411; Roche Diagnostics, Munich, Germany), as previously described [34 (link)]. The serum levels of triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-c), and total proteins will be assessed by colorimetric methods glycerol-phosphate oxidase/peroxidase, cholesterol oxidase/peroxidase, direct detergent, and biuret, respectively. All analyses will be performed using commercially available kits suitable for an A25 automatic analyzer (BioSystems, Barcelona, Spain). Low-density lipoprotein cholesterol (LDL-c) will be calculated using the Friedwald equation [35 (link)].

Maternal weight and height in the third trimester of pregnancy were measured using standard equipment, which allowed us to calculate weight gain and pre-pregnancy body mass index (BMI).
Blood samples were taken from patients after 12 h of fasting. Utilizing an enzymatic colorimetric method using kits, the total cholesterol CHOD-PAP and triglyceride GPO-PAP (Roche Diagnostics GmbH, Mannheim, Germany), serum triglyceride (TG) levels as well as HDL and LDL cholesterol were measured [66 (link),67 (link)]. Glycated hemoglobin (HbA1C) was measured using a latex agglutination test, a turbidimetric immunoassay [68 (link)]. C-reactive protein (CRP) concentration was determined, according to the manufacturer’s instructions, with the use of the cassette COBAS INTEGRA CRP (Latex) in a turbidimetric manner (Roche Diagnostics GmbH, Mannheim, Germany) [69 (link)]. Elecsys insulin assay was used to determine the concentration of plasma insulin (Roche Diagnostics GmbH, Mannheim, Germany) [70 (link)].
For each patient, an indicator of insulin resistance (HOMA-IR) and beta-cell function (HOMA-B) was calculated with the use of the homeostasis model assessment (HOMA) as follows [71 (link)]: $\mathrm{HOMA}-\mathrm{IR}=\mathrm{fasting}\mathrm{insulin}\left(\frac{\mathsf{\mu }\mathrm{U}}{\mathrm{mL}}\right)\ast \mathrm{fasting}\mathrm{glucose}(\frac{\mathrm{mg}}{\mathrm{dL}})/405;$
$\mathrm{HOMA}\mathrm{B}=360\ast \mathrm{fasting}\mathrm{insulin}\left(\frac{\mathsf{\mu }\mathrm{U}}{\mathrm{mL}}\right)/\left(\mathrm{fasting}\mathrm{glucose}\left(\frac{\mathrm{mg}}{\mathrm{dL}}\right)-63\right)$