Background: Chronic ankle instability (CAI) is characterized by pain, repeated sprains and giving way. Recently, the pathomechanical impairment, sensory-perceptual impairment and motor-behavioral impairment have been documented in the CAI model. Previous research revealed that individuals with CAI had lower pressure pain thresholds (PPT) compared to control subjects, which may be related to increased mechanosensitivity of neural tissues around the ankle. Pahor et al. indicated that subjects with repeated ankle sprains showed greater restriction of knee extension during the slump test with ankle plantar flexion and inversion (AKEOST), suggesting restricted mobility of the common peroneal tract. However, the effects of neurodynamic intervention, which addresses the mechanosensitivity problems, in people with CAI are still unclear. Purpose: The aim of the study is to investigate the effects of neurodynamic intervention on the mechanosensitivity, balance performance and self-reported function in patients with CAI. Methods: This is a single-blinded, randomized controlled trial. A total of 36 participants (mean age: 24.64 ± 4.7 years) with CAI were recruited and randomized into either the balance training only group or the neurodynamic intervention with balance training group. Both groups received 12 interventions within 6-8 weeks. Outcome measures included PPT, AKEOST, hamstrings flexibility, ankle ROM, Y balance test and foot and ankle ability measure (FAAM). All data were assessed and collected before and after 12 interventions. Two-way repeated ANOVA was used to examine the treatment effect with the alpha level set at 0.05. Results: There was a significant group by time interaction for AKEOST (p = 0.018) and hamstring flexibility (p = 0.003), favoring neurodynamic intervention. Significant time effects were found for PPT (at the anterior talofibular ligament (p = 0.005), calcaneofibular ligament (p = 0.017), and common peroneal nerve (p = 0.015)), ankle dorsiflexion ROM (p < 0.001), Y-balance test (p < 0.001), and FAAM (p < 0.001). Conclusion: Additional neurodynamic intervention improved the mechanosensitivity and hamstring flexibility in patients with CAI. However, the functional performance outcomes were similar between the two groups. Clinical Relevance: Additional neurodynamic intervention could be integrated in the management plan in patients with CAI.