AIM: The 2017 Japan Atherosclerosis Society (JAS) familial hypercholesterolemia (FH) criteria adopt a cut-off value of ≥ 9 mm of Achilles tendon thickness (ATT) detected by X-ray as one of the three key items. This threshold was determined based on an old data evaluating the ATT of 36 non-FH individuals that was published in 1977. Although the specificity of these clinical criteria is extremely high due to a strict threshold, there are a significant number of patients with FH whose ATT ＜9 mm. We aimed to determine a cut-off value of ATT detected by X-ray to differentiate FH and non-FH based on genetic diagnosis. METHODS: The individuals (male/female=486/501) with full assessments of genetic analyses for FH-genes (LDLR and PCSK9), serum lipids, and ATT detected by X-ray at the Kanazawa University Hospital and National Cerebral and Cardiovascular Center Research Institute were included in this study. Receiver operating characteristic (ROC) analyses were conducted to determine a better cut-off value of ATT that predicts the pathogenic mutation of FH. RESULTS: The ROC analyses revealed that the best cut-off values of ATT are 7.6 mm for male and 7.0 mm for female, with the sensitivities/specificities of 0.83/0.83 for male and 0.86/0.85 for female, respectively. If the thresholds of ATT of 8.0/7.5 mm and 7.5/7.0 mm were applied to the diagnosis of male/female FH, the sensitivities/specificities predicting the pathogenic mutation of FH by the 2017 JAS FH clinical criteria would be 0.82/0.90 and 0.85/0.88, respectively. CONCLUSIONS: These results suggest that the cut-off value of ATT detected by X-ray is obviously lower than 9.0 mm, which was adopted by the 2017 JAS FH clinical criteria.