The binding of human mannose-binding lectin (MBL) to human colon adenocarcinoma cell lines and leukemia cell lines was analyzed by flow cytometry using specific antibodies against MBL. MBL binding was observed in 3 of 7 colon adenocarcinoma cell lines (Colo205, Colo201 and DLD-1), but not in any of 3 leukemia cell lines tested. The binding of MBL to these cell lines was sugar-specific and calcium-dependent, since it was almost completely inhibited in the presence of 10 mM EDTA or 50 mM mannose. The MBL binding to Colo205 cells was more strongly reduced by the pretreatment of the cells with an O-linked glycosylation inhibitor, benzyl-2-acetamide-2-deoxy-alpha-galactopyranoside (Bz-alpha-GalNAc), rather than an N-linked glycosylation inhibitor, tunicamycin. The degree of MBL binding was well correlated with the expression of Lewis A and Lewis B antigens on these cell lines. Moreover, MBL binding to Colo205 cells was inhibited by anti-Lewis A and anti-Lewis B antibodies. These results suggest that MBL could bind to some human colon adenocarcinoma cell lines through their Lewis A and Lewis B moieties.