We have previously shown that inhibition of uPA activity of a human tumor-HEp3-results in a drastic reduction of its metastasis in the chick embryo. Using 125IUdR-labeled tumor cells, we have now studied the role of uPA in individual steps of tumor metastasis. We found that, 48 hr after inoculation of tumor cells on the CAM, the organs of the embryos, inoculated with cells in which uPA was inhibited, contained 4-fold less cells than the controls. Neither the initial advance of the tumor mass into the CAM nor the process of extravasation was affected by the inhibition of tumor uPA. However, the infiltration of the CAM mesenchyme by individual tumor cells was blocked when tumor uPA activity or production was inhibited. In addition, indirect evidence implicated uPA as an essential factor in the tumor cell intravasation.