The monoclonal antibody NCC-CO-450 (IgM kappa) was selected by screening of reactivity with high-molecular-weight antigens (Mr greater than 10(6] isolated from ascitic fluid of a colon cancer patient. This antibody detected heterogeneous but predominantly high-molecular-weight antigens in 4 of 6 ascitic fluid samples from gastrointestinal cancer patients by immunoblotting analysis. A sandwich radioimmunoassay was developed in order to examine the serum level of this antigen, and the cutoff value was defined as the mean plus 2 SD of values obtained with sera from normal donors. While 97% (93 of 96) of sera had a negative antigen value in normal donors, 56% (14 of 25) of patients with colorectal carcinoma and 40% (8 of 20) of patients with gastric carcinoma showed a positive antigen value. The distribution of the antigen in sera of patients with various cancers did not show any correlation with the distribution of carcinoembryonic antigen or CA 19-9. From immunohistochemical and biochemical analyses, NCC-CO-450 antigen was characterized as a mucin-like glycoprotein abundant in normal colonic epithelium as well as in carcinomas of the colon, stomach, and pancreas. The immunohistochemical reactivity of NCC-CO-450 was distinct from that of other monoclonal antibodies reported to be useful for serological diagnosis. The epitope recognized by NCC-CO-450 is considered to be an O-linked carbohydrate chain without terminal sialic acid but is different from the known carbohydrate chains, i.e., Lea, Lex, LeY, Tn, sialyl-Lea, and sialyl sugar chain defined by NCC-ST-439 in a competitive binding inhibition assay of monoclonal antibodies. This newly defined antigen is a good example of a normal antigen shed from cancer cells that can be used successfully as a serum tumor marker.