Step-down heating (SDH) of Chinese hamster ovary cells consisted of an acute heat treatment at an elevated temperature followed immediately by chronic exposure to a lower hyperthermic temperature [step-down temperature (SDT)]. The survival curve at the SDT showed a reduced D0 (dose required to reduce the cell population to 37% on exponential part of the curve) relative to that of control cells only for SDT of 43° C or less. The ratio r of D0's, D0 (control)/D0 (SDH), varied inversely with the SDT (e.g., r = 13 for SDT = 40° C, whereas r = 1.3 for SDT = 43° C). The degree of sensitization, as measured by r, was independent of the conditioning temperature (CT) and the conditioning time, provided that the CT was above 43° C and the conditioning treatment reduced cell survival to 60% or less. The ratio r could be defined mathematically as a function of the activation energies for heat-induced cell killing above and below 43° C, the SDT, and the CT. Biphasic heat survival curves, which appeared with heating at constant temperatures below 43° C, also appeared under SDH conditions, which suggested that thermotolerance, as represented by the resistant phase of the biphasic survival curve, was not affected by SDH. Fractionation experiments to measure the loss of interaction of heat damage remaining from the conditioning treatment with the low-temperature heat damage showed that 63% of the 45° C sublethal damage was removed by 67 minutes at a constant exponential rate.