Our recent studies have shown that the dendritic cell-specific ICAM nonintegrin CD209 (DC-SIGN) specifically binds to the core LPS of Escherichia coli K12 (E. coli), promoting bacterial adherence and phagocytosis. In this current study, we attempted to map the sites within the core LPS that are directly involved in LPS-DC-SIGN interaction. We took advantage of four sets of well-defined core LPS mutants, which are derived from E. coli, Salmonella enterica serovar Typhimurium, Neisseria gonorrhoeae, and Haemophilus ducreyi and determined interaction of each of these four sets with DC-SIGN. Our results demonstrated that N-acetylglucosamine (GlcNAc) sugar residues within the core LPS in these bacteria play an essential role in targeting the DC-SIGN receptor. Our results also imply that DC-SIGN is an innate immune receptor and the interaction of bacterial core LPS and DC-SIGN may represent a primeval interaction between Gram-negative bacteria and host phagocytic cells.