Background: In athletes with anterior cruciate ligament (ACL) deficiencies could assess functional capabilities with different instruments such as use of a camera in vivo situation. However, these methods have suffered from a large number of limitations such as inability to be repeatable and complexity in technique. Objective: The main purpose of this study was to compare gait pattern of the athletes with ACL injury and able-bodied subjects using an accelerometer. Method: A three-dimensional accelerometer was placed over the tibia tuberosity of 20 healthy and 20 individuals with ACL-deficiencies (ACLD). After walking on the treadmill, the principal components of the acceleration data were calculated using MATLAB software. Results: In this study, Principle Component analysis was used for statistical analysis. The results indicated that subjects with ACL deficiency have different gait pattern compared to the control group. The major differences between stride trajectories of the two groups were at the end of mid-swing and the beginning of terminal swing phases in vertical axis. ACL deficient subjects exhibited different gait patterns during mid and terminal stance phases in anterior- posterior axis compared with normal controls. Conclusions: The difference in gait between subjects with ACL deficiency and healthy subjects are depends on variation in the amount of knee flexion and tibia rotation that could be altered to motor recruitment.