Background <p>Chronic Ankle Instability (CAI) is associated with proprioceptive deficits and altered neuromuscular control, yet limited studies have examined these factors concurrently during functional tasks like jump-landing.</p> Methods <p>A total of 44 participants, including 21 healthy individuals and 23 with CAI, participated in a crossover experimental study. An EMG system (Noraxon, USA) was used bilaterally to assess muscle activation of the key muscles involved in ankle movement during jump-landing activities. Specifically, medial gastrocnemius (MG) and tibialis anterior (TA) muscles were evaluated. To identify differences in The Ankle Inversion Discrimination Apparatus for Landing (AIDAL) parameters between groups, an independent samples t-test was conducted. Linear mixed modeling with repeated measures was performed to analyze muscle activation differences during jump-landing at different angles.</p> Results <p>Individuals with CAI exhibited significantly lower AIDAL slope estimates compared to healthy controls, with a moderate effect size (<i>p</i> &lt; 0.001, d = 0.78). In the EMG results, no statistically significant difference was observed between the CAI group and healthy controls (<i>p</i> = 0.431) for TA and (<i>p</i> = 0.699) for MG. Statistically significant differences were found in the activation of the MG muscle between 10^o and 14^o (<i>p</i> = 0.009), between 12^o and 16^o (<i>p</i> = 0.041) and between 14^o and 16^o (<i>p</i> = 0.005).</p> Conclusion <p>This study indicates that individuals with CAI experience impaired proprioception compared to healthy individuals, while showing no notable differences in key muscle activation patterns. It highlights the necessity of rehabilitation strategies focused on proprioception.</p>