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Background: Assessment of calf muscle perfusion requires a physiological challenge. Exercise and cuff-occlusion hyperemia are commonly used methods, but it has been unclear if one is superior to the other. We hypothesized that post-occlusion calf muscle perfusion (Cuff) with pulsed arterial spin labeling (PASL) cardiovascular magnetic resonance (CMR) at 3 Tesla (T) would yield greater perfusion and improved reproducibility compared to exercise hyperemia in studies of peripheral arterial disease (PAD). Methods: Exercise and Cuff cohorts were independently recruited. PAD patients had an ankle brachial index (ABI) between 0.4-0.9. Controls (NL) had no risk factors and ABI 0.9-1.4. Subjects exercised until exhaustion (15 NL-Ex, 15 PAD-Ex) or had a thigh cuff inflated for 5 minutes (12 NL-Cuff, 11 PAD-Cuff). Peak exercise and average cuff (Cuff mean ) perfusion were compared. Six participants underwent both cuff and exercise testing. Reproducibility was tested in 8 Cuff subjects (5 NL, 3 PAD). Results: Controls had greater perfusion than PAD independent of stressor (NL-Ex 74 ± 21 vs. PAD-Ex 43 ± 10, p = 0.01; NL-Cuff mean 109 ± 39 vs. PAD-Cuff mean 34 ± 17 ml/min-100 g, p < 0.001). However, there was no difference between exercise and Cuff mean perfusion within groups (p > 0.6). Results were similar when the same subjects had the 2 stressors performed. Cuff mean had superior reproducibility (Cuff mean ICC 0.98 vs. Exercise ICC 0.87) and area under the receiver operating characteristic curve (Cuff mean 0.992 vs. Exercise 0.905). Conclusions: Cuff hyperemia differentiates PAD patients from controls, as does exercise stress. Cuff mean and exercise calf perfusion values are similar. Cuff occlusion hyperemia has superior reproducibility and thus may be the preferred stressor. [ABSTRACT FROM AUTHOR]