Thrombin immobilized on polystyrene beads initiates DNA synthesis and cell division in quiescent cultures of chick embryo (CE) cells in serum-free medium. These thrombin beads also produce morphological changes in CE cells similar to those produced by soluble thrombin. The amount of acid-precipitable material released from 125I-thrombin-beads into the culture medium was 60-fold less than the amount of soluble thrombin required to produce an equivalent increase in cell number. Moreover, EM autoradiography of CE cells 10 h after 125I-thrombin-bead addition showed that there was no direct release and accumulation of radioactive material in the cytoplasm of these cells. These results demonstrate that thrombin action at the cell surface is sufficient to initiate proliferation of CE cells.