Paracrine motogenic factors, including motility cytokines and extracellular matrix molecules secreted by normal cells, can stimulate metastatic cell invasion. Both intact extracellular matrix molecules and their degradative products may exhibit these activities. We have found that human lung fibroblasts produce paracrine motility-stimulating factors for recently established human sarcoma cell strains. We purified the major fibroblast motility-stimulating factor (FMSF) from human lung fibroblast-conditioned medium by sequential heparin affinity chromatography and DEAE anion exchange chromatography. Lysylendopeptidase C digestion of FMSF and sequencing of peptides purified by reverse-phase high-pressure liquid chromatography identified FMSF as an NH2-terminal fragment of human fibronectin. Using SYN-1 sarcoma cells, FMSF predominantly stimulated chemotaxis and some chemokinesis, and it was chemotactic for a variety of human sarcoma cells, including fibrosarcoma, leiomyosarcoma, liposarcoma, synovial sarcoma, and neurofibrosarcoma cells. The FMSF activity present in human lung fibroblast-conditioned medium was completely eliminated by either neutralization or immunodepletion with a rabbit antihuman-fibronectin antibody, thus further confirming that the NH2-terminal fibronectin fragment was the FMSF responsible for the motility stimulation of human soft tissue sarcoma cells. Because human soft tissue sarcomas have a distinctive hematogenous metastatic pattern (predominantly lung), and lung-derived fibroblasts secrete large amounts of FMSF, FMSF and fibronectin may play a role in stimulating sarcoma invasion into lung tissue.