The Perometer is an optoelectronic device used to calculate limb volume. However, knee angle effect on knee volume during a Perometer measurement has not been reported. This could be a potential confounding factor in assessing knee volume in subjects with impaired knee mobility and where volume measurements are taken over time. To address this issue, bilateral knee volume measurements were recorded from 20 uninjured subjects in seven different knee angles from 0-60 degrees flexion. All 40 knees were not measured in all seven angles because of obstruction of the measurement frame when some knees were in 50 degrees and 60 degrees flexion. Thus, the volume data was separated into three subgroups for analysis: 40 knees at knee flexion angles 0, 10, 20, 30 and 40 degrees; 36 knees at 50 degrees; and 16 knees at 60 degrees. Repeated measures analysis of variance showed a statistically significant difference between the volume measurements in the three subgroups: 0-40 degrees knee flexion [F(4,156) = 35.146, P<0.001], 50 degrees knee flexion (F(5,175) = 56.826, P<0.001] and 60 degrees knee flexion, [F(6,90) = 45.825, P<0.001]. Subsequent paired t-tests showed statistically significant [P<0.001] differences in knee volume for all angle comparisons except for 0 and 10 degrees (absolute volume difference of 1.2 ml; P = 0.772) and 0 and 20 degrees (absolute volume difference of 10.9 ml; P = 0.036) (Bonferroni adjustment applied). The absolute values of the differences that were shown to be statistically significantly different ranged from 12-47 ml for the 0-40 degrees subgroup; 39-85 ml for the 50 degrees subgroup and 35-107 ml for the 60 degrees subgroup. In conclusion, knee angle affects Perometer knee volume measurements and statistically significant differences occur as knee flexion exceeds 20 degrees with greater differences occurring as the knee is more flexed. Hence, knee angle should be reproduced in re-testing and in side-to-side comparisons when evaluating knee volume with the Perometer.