Introduction: Congenital hypothyroidism causes irreversible, irreparable damage and lifelong complications such as mental retardation. Using the Health Belief Model questionnaire, this study aimed to determine the effect of a health education program on mother’s participation for neonatal hypothyroidism screening program. Materials and Methods: In the current interventional study, 46 pregnant women from three Health-medicine centers were randomly assigned to an intervention group and 46 pregnant women from three other health centers were assigned to the control group. The Health Belief Model questionnaire assessed perceived susceptibility, perceived severity, perceived benefits, perceived barriers, and cues to action. For measuring the scientific validity of the designed tool, there was an expert panel, and for reliability we used Cronbach’s Alpha. This questionnaire was completed by both groups before educational intervention. The educational intervention consisted of 4 educational sessions, implemented for the intervention group. After the training, the questionnaire was completed again by both groups. Data analyzed with SPPS software using χ² and ANCOVA. Results: There were statistically significant deferencse between the two groups regarding the perceived susceptibility, severity, benefits and cues to action scores after the training sessions (p<0.001). The difference in mean barrier scores of the two groups was not statistically significant after the training sessions (p=0.16). Timely referral by mothers for neonatal hypothyroidism screening was higher in the intervention group (p<0.001). Conclusion: The study indicated that educational programs based on Health Belief Model assessment are effective in timely participation of mothers for neonatal screening and its use is strongly recommended in screening programs.