Objectives The 6-minute stepper test (6MST) is a submaximal test that requires little space to assess exercise capacity compared to the 6-minute walk test (6MWT). The study aims to investigate the test-retest reliability and convergent validity of 6MST and to compare physiological responses, dyspnea, fatigue perception with 6MST and 6MWT in patients with ankylosing spondylitis (AS). Methods To test the convergent validity of 6MST, 65 patients performed both 6MWT and 6MST on the first day and correlation between two tests were assessed with Pearson correlation test. In order to investigate the test-retest reliability of the 6MST, 32 of the 65 patients performed 6MST one week later and intraclass correlation coefficients (ICC) were calculated. Dyspnea and fatigue perception were analyzed with using Wilcoxon signed-rank test, physiological responses were analyzed using paired sample t-test. Results Excellent test-retest reliability was observed for 6MST (ICC: 0.988). There was a significant correlation between 6MST and 6MWT (r: 0.725, p < 0.001). Dyspnea and leg fatigue perception were significantly higher in 6MST (p < 0.05). Physiological responses and fatigue perception were similar in both 6MST and 6MWT (p > 0.05). Conclusion This study demonstrated that the 6MST is reliable and valid method to evaluate exercise capacity in patients with AS. 6MST can be used to evaluate exercise capacity of patients with AS.