Blood samples were obtained by venipuncture, refrigerated immediately, and transported to the central testing institute. Fasting plasma glucose and serum lipids (total cholesterol, high-density lipoprotein cholesterol, and triglycerides) were measured enzymatically with the Hitachi automated analyzer 7600 (Hitachi, Tokyo, Japan), and fasting plasma insulin was determined using a 1470 WIZARD gamma-Counter (Perkin-Elmer, Turku, Finland) and an immunoradiometric assay (Biosource, Fleurus, Belgium). The homeostasis model assessment of insulin resistance (HOMA-IR) was calculated as a marker of insulin resistance, as follows:
The 25-hydroxyvitamin D concentration was measured by radioimmunoassay using a 1470 WIZARD gamma-Counter (Perkin-Elmer) and a 25-hydroxyvitamin D 125I radioimmunoassay kit (DiaSorin, Stillwater, MN, USA). At serum 25-hydroxyvitamin D levels of 8.6, 22.8, 33.1, and 49.1 ng/mL, the inter-assay coefficients of variation (CV) were 11.7%, 10.5%, 8.6%, and 12.5%, respectively, and the intra-assay CVs were 9.4%, 8.2%, 9.1%, and 11.0%, respectively.
The 25-hydroxyvitamin D concentration was measured by radioimmunoassay using a 1470 WIZARD gamma-Counter (Perkin-Elmer) and a 25-hydroxyvitamin D 125I radioimmunoassay kit (DiaSorin, Stillwater, MN, USA). At serum 25-hydroxyvitamin D levels of 8.6, 22.8, 33.1, and 49.1 ng/mL, the inter-assay coefficients of variation (CV) were 11.7%, 10.5%, 8.6%, and 12.5%, respectively, and the intra-assay CVs were 9.4%, 8.2%, 9.1%, and 11.0%, respectively.