Human monocytes bind lipopolysaccharides (LPS) complexed to LPS binding serum proteins (LBP; septin) via surface glycoprotein CD14. Flow cytometry was used for the measurement of endotoxin binding to monocytes using fluorescein isothiocyanate (FITC)-labeled LPS. LPS-FITC binding to bovine monocytes was mediated by calf serum. The addition of purified human soluble CD14 (sCD14) to the calf serum abrogated the binding of LPS-FITC to bovine monocytes. The function of sCD14 could specifically be blocked by a monoclonal antibody (MEM-18) competing for LBP-LPS binding to human CD14 but not by a noncompeting antihuman CD14 monoclonal antibody (mAb) (RoMo-1) or irrelevant antibodies. Both mAbs MEM-18 and RoMo-1 are restricted to the human system. Concerning the species specifity of monoclonal antihuman CD14 antibodies, it is possible to demonstrate the specific function of purified human soluble CD14 for neutralization of endotoxin using the bovine system. These results demonstrate the biological function of the serum protein sCD14 protecting cells from endotoxin-induced cell activation, which becomes relevant for pathological situations such as ongoing septic shock. The LBP-LPS binding region of CD14 glycoprotein seems to be a highly conserved structure similar in different species.