Serum is known to be required for invasion or phagokinesis of certain tumor cells, although the mechanism of its action is not well understood yet. In the in vitro invasion assay system we have developed, MM1 cells exhibiting extensive invasiveness against cultured mesothelial cell monolayers in the presence of 10% fetal calf serum did not invade them without serum. Human small cell lung cancer (OC10) cells also required serum in invasion. Serum could be completely substituted by oleoyl-lysophosphatidic acid (LPA) or bacterial phospholipase D (PLD). Phosphatidic acid also had an invasion-inducing activity, though to a lesser degree. Since human ovarian cancer (RMUG-S) cells require neither serum, LPA nor the PLD for invasion of mesothelial cell monolayers, the production of phospholipid like LPA, a candidate for intracellular second messenger, in tumor cells seems critical for the invasion by MM1 cells or OC10 cells. This result suggests a possible participation of particular signaling cascade, PLD-LPA(PA) system, in the invasion of certain tumor cells.