Elisa solid phase direct sandwich method

The CD4 lymphocytes were measured by cytometry technique using a flux cytometry (Apogee Flow systems Limited® Apogee Flow Systems, Hertfordshire, United Kingdom). High sensitivity C-reactive protein (hs-CRP) was measured using the ELISA solid-phase direct sandwich method (Sigma-Aldrich, St. Louis, USA) with ELx808™ Microplate reader (BioTek Instruments, Winooski, USA).

Venous blood was collected after overnight fasting to determine fasting plasma glucose (FPG) using Accu-Chek® Active reader, as described by the manufacturer (Roche Diabetes Care GmbH, Sandhofer Strasse 116, 68305 Mannheim, Germany). Plasma concentrations of total cholesterol and triglycerides (TG) were evaluated by enzymatic methods. High-density lipoprotein (HDL) cholesterol was evaluated by enzymatic colorimetric method. Low-density lipoprotein (LDL) cholesterol was calculated using the Friedewald formula if the triglycerides are less than 400 mg/dl (4.6 mmol/l) [16 (link)]. Biochemical tests were performed using a MaestroNano® Pro Spectrophotometer (Maestrogen, 8275 South Eastern, Avenue #200, Las Vegas, NV 89123, USA) and the reference commercial kits (Sigma-Aldrich Co., 3050 Spruce Street, St. Louis, MO 63103, USA). We analyzed high sensitivity C-reactive protein (hs-CRP) using ELISA solid phase direct sandwich method (Sigma-Aldrich, St. Louis, USA) with ELx808™ Microplate reader (BioTek Instruments, Winooski, USA).