We have reported previously that the production of a tumor cell factor that stimulates synthesis of fibroblast collagenase is influenced by a fibroblastdeposited matrix component, possibly heparan sulfate-proteoglycan. In this study, binding sites for heparin and heparan sulfate on mouse 6-1 6 melanoma cells have been demonstrated. Binding of 'H-heparin and "S-heparan sulfate has been shown to occur to whole cells, isolated membranes, and to a component(s) of detergent extracts of the membranes. Scatchard analysis of binding of 'H-heparin yielded a Kd of 2-5 x M and a B, , , , of 0.5 x lo7 heparin molecules bound per cell. Binding of "5-heparan sulfate was of at least an order of magnitude lower affinity than heparin, but the B , , , was similar to that for heparin. Competition studies showed that 35S-heparan sulfate binding was inhibited totally by heparin and heparan sulfate and partially by dermatan sulfate, but no inhibition was obtained with hyaluronate or chondroitin sulfate. Binding of 'H-heparin was inhibited totally by heparin but to different extents by preparations of heparan sulfate from different tissue sources. The hepariniheparan sulfate binding activity is a protein(s) because it is destroyed by treatment with trypsin. Binding of 3H-heparin to transblots of the detergent extract of the 6-16 cell membranes indicated that at least part of the binding activity is a 14,000dalton protein