Whole blood and peripheral blood mononuclear cell (PBMC) culture models have been used to study cytokine stimulation and release in vitro. In this study, we characterize the kinetics of the interleukins (IL-1 beta), (IL-6), (IL-8), and tumor necrosis factor-alpha (TNF-alpha) following an endotoxin (ET) challenge using our in vitro whole blood model. Whole blood samples from 10 healthy volunteers were studied. All cytokines were measured by enzyme-linked immunosorbent assay. Peak concentrations of TNF-alpha occurred 2 h after ET challenge followed by a rapid decline in free plasma TNF-alpha concentration (half-life 18.2 min). IL-1 beta was not significantly elevated until 4 h after ET challenge. IL-8 was elevated 1 h after ET challenge. IL-6 concentration exhibited a biphasic peak occurring at 6 and 74 h after ET challenge. We conclude that (1) our whole blood in vitro model produces cytokine release kinetics similar to those reported in vivo, and (2) the presence of either binding proteins or cellular metabolism of TNF-alpha in whole blood produces a similar plasma half-life to that observed in vivo.