Introduction: In Brazil, the law 13.002/2014 states that all newborns in the country must undergo lingual frenulum assessment before hospital discharge. Objective: To validate the Lingual Frenulum Protocol for Infants. Material and Methods: The study used data from 100 infants assessed at 30 days of life at Santa Therezinha Hospital, in Brotas, SP, Brazil. The LFPI was administered to the 30-day-old infants and all assessments were registered in patient records and videotaped. Live exams, assessments and re-assessments using images of the infants were performed by two examiners experienced in Orofacial Myofunctional Disorders. The study included healthy full-term infants of both genders, who were being breastfed exclusively. Prematurity, perinatal complications, craniofacial anomalies, neurological disorders, and visible genetics syndromes were the exclusion criteria. The validation process included the analysis of content validity, criterion validity, construct validity, reliability, sensitivity, specificity, positive predictive value, and negative predictive value. Results: Concerning content validity, the percentage of agreement was 100%. The criterion validity was measured by comparing the common items of the LFPI and BTAT. The agreement percentage for the analyzed items was 100%. The construct validity was analyzed by comparing the partial and total scores from the assessment of infants with and without lingual frenulum alteration at 30 and 75 days of life. Results demonstrated similarity of scores between the control group and the group that underwent frenotomy. The results from the inter-rater and intra-rater agreement analysis indicated perfect agreement. The values of sensitivity, specificity, positive predictive value, and negative predictive value were 100%. Conclusion: The study concluded that the LFPI was a valid and reliable assessment too that ensures accuracy when diagnosing lingual frenulum alterations within the investigated parameters.