We provide several independent lines of evidence linking microneme secretion and Toxoplasma invasion of host cells based on: (i) temperature dependence; (ii) inhibition by temperature cycling; (iii) the requirement for intracellular calcium; and (iv) inhibition by staurosporine. Additionally, our inhibitor studies indicated that microneme exocytosis is controlled by a calcium-dependent pathway involving protein phosphorylation. Furthermore, we observed a dramatic redistribution of surface MIC2 during parasite penetration into the host cell. Collectively, these studies indicate that secretion of micronemal proteins is tightly associated with Toxoplasma invasion of host cells, which is consistent with our hypothesisthat micronemal proteins play animportant role in host cell invasion.