P-FPMI-170 - P-FPMI-170
Venous blood (20 ml) from healthy volunteers was collected in Vacutainer® collection tubes containing sodium heparin as an anticoagulant (Becton Dickinson, Mississauga, ON) in accordance with UBC ethical approval and guidelines. Blood was diluted 1:1 with complete RPMI 1640 medium and separated by centrifugation over a Ficoll-Paque® Plus (Amersham Biosciences, Piscataway, NJ, USA) density gradient. White blood cells were isolated from the buffy coat, washed twice in RPMI 1640 complete medium, and the number of peripheral blood mononuclear cells (PBMC) was determined by trypan blue exclusion. PBMC (4 x 106) were seeded into 6-well tissue culture dishes (Falcon; Becton Dickinson) at 1x106 cells/ml at 37°C in 5% CO2. Cells were treated with LPS (purified from Pseudomonas aeruginosa), at 2ng/ml for 4 hrs.