Enzymatic reagents

Blood draws were performed at the time of the baseline visit following an overnight fast. Blood samples were allowed to clot at room temperature for 30 minutes, and serum was separated by centrifugation at 1,500 g at 4 °C for 20 minutes. Aliquots were stored at −80° C in the MOST repository. For the determination of lipid profiles, matched case-control samples (N=994) were shipped overnight on dry ice to the Cardiovascular Nutrition Laboratory at the Jean Mayer USDA Human Nutrition Research Center of Aging at Tufts University. Serum total cholesterol and HDL concentrations were measured on an AU400e automated analyzer (Beckman Coulter, Brea, CA; intra-assay CV< 3%; inter-assay CV <4%) using enzymatic reagents (Beckman-Coulter). LDL concentration was calculated using the Friedewald equation (19 (link)) except when triglycerides were above 400mg/dl. For those samples, a direct LDL method was used (AU400e automated analyzer, Beckman Coulter, Brea, CA; intra-assay CV< 2.4%; inter-assay CV <3.6%).

Serum concentrations of total cholesterol and low-density lipoprotein cholesterol (LDL-C) were measured using an AU400 clinical chemistry analyzer with enzymatic reagents (Beckman Coulter, Inc.) at the Jean Mayer USDA Human Nutrition Research Center on Aging at Tufts University, Boston, MA. Fasting plasma glucose concentrations were measured by One Touch® glucometer. Fasting insulin concentrations were determined by ELISA using a commercial kit (Millipore, Billerica, MA). The insulin resistance was evaluated using homeostatic model assessment for insulin resistance (HOMA-IR) by the following formula:
$$\text{HOMA-IR}=\left[\text{Fasting plasma glucose }\left(\text{mg}/\text{dL}\right)\times \text{Fasting plasma insulin }\left(\text{mU}/\text{L}\right)\right]\times {\left(405\right)}^{-1}$$