There has been considerable interest in the effect that gram-negative bacterial endotoxin (lipopolysaccharide [LPS]) can have in asthma, given that inhalation of LPS has been shown to cause bronchial hyperresponsiveness. Further, there is evidence that the endotoxin-binding protein CD14 may be a marker for asthma. Inhaled LPS has been shown to cause an influx of eosinophils into the nasal airway and to increase the survival of CD16-negatively selected eosinophils in vitro. In this study, we compared survival of eosinophils isolated via CD16-negative selection with eosinophils that were isolated using both CD16- and CD14-negative selection criteria. Survival of CD16-negatively selected eosinophils was enhanced by LPS in a dose-dependent manner and was inhibited by the endotoxin antagonists polymyxin B or lipid X. In contrast, depletion of CD14(+) cells within the eosinophil preparations (CD14/CD16-negatively selected eosinophils) decreased the effect of LPS on survival. Preincubation of CD16-negatively selected eosinophils with antibody 60bd, which blocks LPS binding to CD14, prevented the survival-enhancing effect of LPS. However, CD14 was not detected on eosinophils by flow cytometry, even after incubation with LPS for up to 24 h. These results suggest that the survival-enhancing effect of LPS on eosinophils requires the presence of CD14(+) cells in the population. It is our hypothesis that enhanced eosinophil survival with LPS involves the contribution of another cell type.