Objective: This double-blind randomized 12-leg cross designed study aimed to investigate the effect of three post-brushing regimens on the salivary fluoride concentration. Methods: Eighteen young healthy volunteers aged between 4.5 ± 0.1 years were recruited to participate in this study. The experiments were performed day other day for 24 days. Before starting the experiment, baseline whole mixed saliva samples were collected. All subjects were randomly brushed with 500 and 1000 ppm sodium fluoride toothpaste with two different amounts: smear-sized (0.13 g) and pea-sized (0.25 g). After one-min tooth brushing by trained caretakers, three post-brushing activities: (1) rinsing with 10 ml de-ioninized distilled water for 10 s, (2) wiping with cleaned cloth, and (3) just splitting out were randomly assigned to each subject. After completing each experimental procedure, whole mixed unstimulated saliva samples were collected at different time intervals up to 45 min for fluoride analyses. Results: The baseline salivary fluoride concentration (mean ± S.D.) was 0.058 ± 0.002 ppmF. During the first 10 min. after tooth brushing, the salivary fluoride concentration of all groups was obviously increased (≥ 0.08 ppm F) and slowly returned to the baseline level after 45 min. Tooth brushing with pea-sized toothpaste containing 1000 ppm F resulted in the highest levels of salivary fluoride retention. However, there was no statistically significant difference in the salivary fluoride concentration among three post-brushing regimens. Conclusion: Salivary fluoride retention related to the amount and concentration of fluoride toothpastes, but not the post-brushing behaviors performed in this study.