BACKGROUND: An ankle sprain is a relevant public health issue. Fatigue changes the neuromuscular response in people with chronic ankle instability (CAI). AIM: To evaluate the muscle co-activation on people with chronic ankle instability using cross-correlation analysis. METHOD: Twenty-four healthy women were selected and divided into stability and instability groups. Ankle sprain was simulated with a mechanical platform. Electrical muscle activity (fibularis brevis, FB; fibularis longus, FL; gastrocnemius lateralis, GL; and tibialis anterior, TA) and platform acceleration were recorded at 2KHz. Two sets of 8 right and eight left foot fall in random order were performed before and after the fatigue protocol. Fatigue protocol ended when the volunteer increased the test run time by 150% of the best round. Co-activation was calculated with cross-correlation. Agonist-agonist (FB-FL, FB-GL, and FL-GL) and agonist-antagonist (TA-GL, TA-FB, and TA-FL) pairs were evaluated. Statistical significance was p<0.05. RESULTS: Co-activation was lower for the instability group. Fatigue did not induce changes in 5 out of the 6 analyzed muscle pairs. CONCLUSION: CAI is a factor of joint instability. Fatigue may not be relevant in altering joint stability. Therefore, interventions should be focused on enhancing joint stability.