Prestimulation with recombinant human thyroid-stimulating hormone (rhTSH) augments radioiodine 131 I therapy for benign nontoxic multinodular goiter. The purpose of this study was to determine the optimal time interval between rhTSH and 131 I administration to enhance thyroid radioactive iodine uptake (RAIU). Methods: Patients were randomized, in a 2-factorial design, to receive either a 0.1-mg dose of rhTSH (n 5 60) or placebo (n 5 30) and to a time interval of 24, 48, or 72 h before 131 I administration. The rhTSH-or placebo-stimulated RAIU study was performed at 4 wk after a baseline RAIU assessment in a tertiary referral center at a university hospital. A total of 90 patients (78 women; median age, 52 y; range, 22-83 y) referred to 131 I therapy for symptomatic nontoxic goiter (median goiter volume, 63 mL; range, 25-464 mL) were included in the study. Change in thyroid RAIU was determined at 24 and 96 h after 131 I tracer administration. Results: In the placebo subgroups, RAIU did not change significantly from baseline. The mean (6SE) 24-h RAIU increased from 33.8% 6 2.3% to 66.0% 6 1.8% (111.2% increase) with a 24-h interval, from 36.8% 6 2.1% to 64.6% 6 2.7% (83.3% increase) with a 48-h interval, and from 33.0% 6 2.7% to 49.6% 6 2.5% (62.4% increase) with a 72-h interval. All within-group changes were highly significant (P , 0.001). The effect was negatively correlated with initial RAIU (r 5 20.703, P , 0.001). The increase in 24-and 96-h RAIU was significantly higher in the rhTSH/24-h group than it was in the rhTSH/72-h group (P 5 0.023 and 0.012, respectively) and insignificantly higher than in the rhTSH/48-h group (P 5 0.37 and 0.26, respectively). Conclusion: The effect of rhTSH on thyroid RAIU is most pronounced when administered 24 h before 131 I administration and declines with longer time intervals. Whether there is a similar time dependency for goiter reduction after rhTSH-stimulated 131 I-therapy remains to be clarified.