Three NIH3T3 transformants, MTAg, MTPy, and MT1, which grow similarly in soft agar media, showed remarkable differences in athymic mice: MTAg grew more rapidly than MTPy, whereas MT1 and NIH3T3 did not, and only MTAg metastasized in lung. Structural analysis of N-glycans from plasma membrane glycoproteins revealed that each sample contains similar amounts of N-glycans, but the relative amounts of 2,6-branched tri- and tetra-antennary oligosaccharides prominently increase and the relative amounts of biantennary oligosaccharides prominently decrease in the order of NIH3T3, MT1, MTPy, and MTAg, whereas those of others remained constant. Western blot analysis revealed that binding of Datura stramonium agglutinin, which interacts with 2,6-branched tri- and tetra-antennary oligosaccharides, is significantly increased in several bands from MTAg compared with NIH3T3, two of which are tentatively identified as lysosome-associated membrane protein-1 and fibronectin (FN)-receptor. It was also shown that the spreading of MTAg on FN-coated plates is dramatically inhibited with the anti-FN-receptor antiserum when compared with NIH3T3. These results indicate that the increased expression of highly branched N-glycans at cell surface is correlated with the rapidness of tumor formation and altered adhesive properties of tumor cells in vivo.