Only physiological levels of melatonin exert an antiproliferative effect on MCF-7 breast cancer cells grown in an anchorage-dependent culture system. We investigated melatonin's effect on the anchorage-independent growth of MCF-7 cells as well as the dose-response characteristics of this indoleamine under clonogenic culture conditions. Melatonin's inhibitory effect, with respect to the number and size of colonies formed, exhibit a linear dose-response curve with pharmacological concentrations producing a maximal inhibition while subphysiological levels of melatonin induce minimal inhibition. These results indicate that cellular attachment may modify the sensitivity of MCF-7 cells towards melatonin.