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Background: Patellofemoral pain syndrome (PFPS) is one of the common causes of anterior knee pain in young people.Objectives: This study investigated the length difference of lower limbs in patients with PFPS syndrome.Methods: In this descriptive cross-sectional study, the difference in the length of the lower limbs of patients in the supine position was assessed using five different clinical methods. These five methods included: 1- the measurements of the actual length of the lower limb, 2- the apparent length of the lower limb, 3- the method of Iliac Crest Palpation and Book Correction (ICPBC) under the short leg of the patient, 4- evaluation of the internal ankles, and 5- bilateral anterior superior iliac spine (ASIS). Data were analyzed using the SPSS software.Results: Nearly 60% of the patients with PFPS felt pain in both knees. The average actual length difference of the lower limbs in patients was 7.00±4.04 mm. Moreover, 75% of the patients had a limb length difference of more than 4 mm. There was no significant difference in the actual length of the limbs regarding age, BMI, employment status, and history of underlying diseases. However, these factors were significantly higher among women than men (p=0.047). Moreover, the results showed that the average length of limbs was significantly lower among patients who exercised during the week (p=0.015). Measurement of lower limb length had moderately correlated using two methods of actual limb length and ICPBC (p=0.001, Kappa=0.421) and two methods of apparent limb length and medial ankle (p=0.032, Kappa=0.317).Conclusions: The difference in lower limb length was more than 4 mm in most patients (75%). Furthermore, in most patients with knee pain on the left side (unilateral/bilateral left more severe), their right side was short and vice versa.