The lethal effects of hydroxyurea (HU) and 3H-thymidine (3H-dT) on mouse hematopoietic cells were compared after various experimental procedures. The aim was to explore the relative efficiency of these 2 methods in analyzing the kinetic properties of progenitor cells. Both methods indicated that 40-50% of progenitor cells assayed with diffusion chamber culture (DCPC) were in S phase 3 days after cyclophosphamide treatment. Effects of HU, but not 3H-dT, were altered by neostigmine triggering of normal DCPC into cell cycle. Cooling marrow cells before exposure to HU or 3H-dT largely abrogated the effect of HU, but not of 3H-dT. Blood-borne DCPC were not in cycle according to the HU effect. Separated blood DCPC were apparently in cycle, as judged with 3H-dT, but the Isopaque-Ficoll separation procedure rendered normal marrow DCPC susceptible to 3H-dT killing. When marrow cells were cultured in DC, the HU technique appeared to be suitable for evaluation of modulation of progenitor cell (CFU-S or CFU-C) proliferation, but previous experiments showed that the 3H-dT technique is a convenient method to assess the initial triggering of CFU-S into cycle in DC culture.