Access 2 immunoassay system

Nonfasting serum was stored at 4 °C and was used for lipid profile assays within 12 hours after sampling. Total cholesterol (TC), TG, LDL-c, HDL-c, and sd-LDL-c were measured utilizing commercial kits using enzymatic methods on an automatic biochemical analyzer (Siemens ADVIA 2400 Chemistry System). The remnant cholesterol was determined using a commercial enzymatic assay (Shanghai Runho Biotech Ltd., China). The assay principle could be explained by the fact that surfactants cooperating with phospholipase D exhibited favorable selectivity towards remnant lipoproteins, making their component cholesterol available for enzymatic assays. The assay was validated to be equivalent to the RemL-C Kit (Kyowa Medex Co., Ltd., Japan) with a correlation coefficient of 0.98 and an interassay CV of 1.8% in our laboratory. The measurement of cardiac troponin I (cTnI), creatine kinase (CK-MB), and myoglobin (MYO) was performed using commercial chemiluminescence immunoassays on an automatic immunoassay analyzer (Beckman Coulter Access 2 Immunoassay System). Non-HDL-c was calculated as TC minus HDL-c, and the AL-c/HDL-c ratio was calculated as the sum of sd-LDL-c and RLP-c divided by HDL-c. Other data were collected by reviewing the medical records of patients.

Serum samples were used for estimation of IgE and Vitamin D levels on Access 2 Immunoassay System (Beckman Coulter, Inc., USA) using CLIA technique. Whole blood samples were used for estimation of eosinophil count on Sysmex XP-300™ Automated Hematology Analyzer (Sysmex America, Inc., USA) and Erythrocyte sedimentation rate (ESR) using Westergren’ method. IgE level of > 180 IU/ml was taken as “Elevated”. In case of Vitamin D levels, < 20 ng/ml was considered as “Deficient”, 20–30 ng/ml as “Insufficient”, 31–100 ng/ml as “Sufficient” and > 100 ng/ml as “Upper safety limit”. Relative eosinophil count > 6% of peripheral blood was considered as “Elevated”. ESR of > 15 mm/hr in males; > 20 mm/hr in females; > 10 mm/hr in children was taken as “Elevated”.

Total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), fasting plasma glucose (FPG) and FTBAs were tested using Dimension RxL auto analyzer (Dade Behring Diagnostics, Deer-field, IL, USA). Glycated hemoglobin A1c (HbA1c) was estimated by high-performance liquid chromatography using HLC-723G7 analyzer (Tosoh Corporation, Tokyo, Japan). Fasting serum insulin (FINS) and C-peptide were measured by Access 2 immunoassay system (Beckman Coulter, Inc., Brea, CA, USA). FTBAs was measured by Total Bile Acids Kit (Enzymatic Cycling assay) (Biosino Bio-Technology and science Inc., Beijing, China). Homoeostasis model assessment for insulin resistance (HOMA-IR) = FPG (mmol/L) × FINS(mU/L)/22.5, and homeostasis model assessment β (HOMA-B) was used to estimate β-cell function and calculated by HOMA-B = 20 × FINS (mU/L) / [FPG (mmol/L) − 3.5] [17 (link)]. Changes in parameters were expressed as delta parameters, Δ = parameter (after exenatide treatment) − parameter (pre-treatment).

Serum albumin-adjusted Ca and phosphate levels were measured in the laboratory by standard methods using the Roche Diagnostics Modular Analytic system (Roche Diagnostics, Indianapolis, IN). PTH level was measured by an Access 2 immunoassay system (Beckman Coulter, Brea, CA), and the inter- and intra-assay CVs were 5.8% and 4.5%, respectively. The normal range for serum PTH level was accepted to be between 12-88 pg/mL.

The following biochemical markers were measured in the serum: serum glucose, total cholesterol and its fractions of low-density lipoprotein (LDL), high-density lipoprotein (HDL), and very low-density lipoprotein (VLDL). The other markers were lipoprotein A (LPA), apolipoprotein A1 (apoA1), apolipoprotein B (apo B), Urea, Creatinine, Calcium, Phosphorus, Uric acid, Alkaline phosphatase, and high sensitivity C-reactive protein. The biochemical parameters were measured using the COBAS INTEGRA® 400 plus (Roche), according to the manufacturer's protocols.
The hormonal markers: cortisol, vitamin D, 17-Beta Estradiol, beta estradiol, follicle stimulating hormone (FSH), luteinizing hormone (LH), thyroid stimulating hormone (TSH), total testosterone, sex hormone binding globulin (SHBG), dehydroepiandrosterone sulfate (DHEAS) and insulin were measured using the ACCESS 2 IMMUNOASSAY SYSTEM® (Beckman Coulter).