Anti-CD14 antibody can inhibit the lipopolysaccharide (LPS)-induced systemic inflammatory response syndrome in case of bacteremia or endotoxemia. To obtain chimeric anti-CD14 antibody, we constructed and expressed a novel chimeric antibody Hm2F9 composed of anti-CD14 single-chain fragment variable (scFv) and the Fc region (the hinge, CH2, and CH3 domains) of human IgG1 in Chinese hamster ovary (CHO) cells based on our previous study of scFv2F9. The Hm2F9 antibody, sized 150 kDa, retained the strong specific antigen-binding ability to the CD14 antigen with a comparable activity (the percentage of positive cells 99.07%) to its parental murine antibody 2F9 (the percentage of positive cells 98.86%). At the same time, Hm2F9 could manifestly block the binding of LPS to CD14, whose positive-cell percentage drops significantly with percentage of 98.63% (from 98.37% to 1.35%). The chimeric antibody Hm2F9 expressed in CHO cells retained high affinity to human CD14 and biological function to LPS.