ObjectivesPopliteal artery entrapment syndrome (PAES) is a rare condition where musculoskeletal structures compress the popliteal artery (POPA) leading to vascular compromise. This study investigates the effect of dynamic plantar‐ and dorsi‐flexion loading on POPA hemodynamic parameters to develop a robust diagnostic ultrasound‐based protocol for diagnosing functional PAES.MethodsHealthy individuals (n = 20), recreational athletes (n = 20), and symptomatic (n = 20) PAES patients were consented. Triplex ultrasound imaging of lower limb arteries was performed (n = 120 limbs). Proximal and distal POPA's in dorsi‐/plantar‐flexion, in prone and erect positions, were imaged at rest and flexion. Peak systolic velocities (cm/s) and vessel diameter (antero‐posterior, cm) was measured.ResultsDistal vessel occlusion was noted across all three groups whilst prone during plantar‐flexion (62.7%). POPA occlusion was only noted in the proximal vessel within the patient group (15.8%). When prone, 50% of control (n = 40 limbs), 70% of athletes (n = 40 limbs), and 65% of patients (n = 40 limbs) had distal POPA occlusion in plantar‐flexion. When prone, recreational athletes (5%), and patients (12.5%) had distal POPA compression under dorsi‐flexion. POPA occlusions with the patient in erect position were only noted in the symptomatic patient group under both dorsi‐flexion (15.8%) and plantar‐flexion (23.7%).ConclusionCompression of the POPA on ultrasound should not be the sole diagnostic criteria for PAES. POPA compression exists in asymptomatic individuals, primarily under prone plantar‐flexion. To reduce false positives, ultrasound‐based protocols should focus on scanning patients in the erect position only to diagnose PAES, rather than asymptomatic POPA compression. A distinction should be made between the two.