Human blood sample preparation
Venous blood was collected from healthy volunteers using EDTA vacutainers (Becton Dickinson, Franklin Lakes, NJ) following informed consent. The study was approved by the University of Louisville Institutional Review Board (08.018). Volunteers were excluded for the presence of chronic illness, use of anti-inflammatory or immunosuppressive medications or signs of acute illness. Whole blood was aliquoted in 1 mL volumes in Falcon polypropylene culture tubes (Fisher Scientific, Waltham, MA). Samples were incubated for up to 18 hours in various conditions at 37°C with 5% carbon dioxide and gentle vortexing every 4 hours. After 18 h, whole blood was centrifuged at 3000 RPM for 3 minutes to obtain plasma which was stored at -80°C until later analysis. Prior work determined optimal sample timing [35].