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Background: Prevalence of ankyloglossia may vary depending on the assessment tool. This systematic review aimed to evaluate the prevalence of ankyloglossia in distinct age groups according to different assessment tools., Types of Studies Reviewed: Nine electronic databases were searched from inception through November 2021 without restrictions of language or year of publication. Paired independent reviewers selected cross-sectional and cohort studies reporting the diagnosis of ankyloglossia, extracted data, and assessed methodological quality. The number of patients with ankyloglossia and the sample were extracted to calculate the overall prevalence of ankyloglossia and 95% CI. The authors calculated the prevalence of ankyloglossia per assessment tool, age group, and sex. They assessed the certainty of evidence using the Grading of Recommendations Assessment, Development and Evaluation approach., Results: Seventy-one studies were included. Seven different diagnostic tools were used. The overall prevalence of ankyloglossia was 5% (95% CI, 4.0% to 5.0%) and ranged from 2% (using an unspecific tool) to 20% (Coryllos classification). The prevalence per age group was higher in infants (7%). The prevalence ratio was 1.34 (95% CI, 1.17 to 1.54) for boys, with very low certainty of evidence., Practical Implications: The prevalence of ankyloglossia is higher among infants and differs depending on the assessment tool used for the diagnosis. It is uncertain whether boys are more affected by ankyloglossia than girls., (Copyright © 2022 American Dental Association. Published by Elsevier Inc. All rights reserved.)