“…Peripheral blood mononuclear cells were prepared by Ficoll separation of peripheral blood from healthy volunteers as previously described [ 13 ]. Mononuclear cells were plated at a density of 10 6 cells/mL in 1.5 mL RPMI 1640 (Irvine Scientific, Santa Ana, Calif), supplemented with 100 U/mL penicillin/streptomicin, 2 mM glutamine, and 3% fetal calf serum, and treated with PepG (Sigma, St Louis, Mich), bactDNA (KM Biomedicals, Aurora, Ohio; LPS < 0.2 EU/mL measured by the limulus amebocyte lysate assay [ 13 ]), or both. TNF- α was measured by ELISA from the culture supernatant after 5-hour incubation and elimination of nonadherent cells.…”