In this study, cell proliferation was examined at low and high cell densities, using HHUA and Jurkat cell lines as experimental models for the antiproliferative and proliferation-enhancing effects of GnRH agonist, Buserelin, respectively. For efficient evaluation of Buserelin activity at low cell density, the colony-forming efficiency assay was adopted. Buserelin markedly affected colony-forming efficiency in a dose-dependent manner at low cell density; however, Buserelin had no effect at high cell density. The conditioned medium of HHUA cells inhibited the Buserelin action, whereas that of Jurkat cells mimicked it. These results suggest that each cell line secretes some substances which regulate cell proliferation, and that these substances can also change the effects of Buserelin. The measurement of colony-forming efficiency is a very effective way of eliminating autocrine and/or paracrine effects, and is a highly sensitive method for measuring GnRH activity.