Purpose: To evaluate the retinal nerve fiber layer (RNFL) thickness of eyes with amblyopia using optical coherence tomography (OCT). Methods: RNFL thickness measurements obtained by time-domain OCT of amblyopic eyes of patients with strabismus (35 patients) and patients with anisometropia (30 patients) were compared with their fellow eyes and age- and gender-matched healthy eyes (40 participants). RNFL thickness values of the groups were also compared and the correlation between RNFL thickness and refraction was evaluated. Results: In the strabismic and anisometropic groups, there were no statistically significant diff erences in average RNFL thickness between amblyopic eyes and their fellow eyes (P > .05). Temporal RNFL quadrant in the hypermetropic anisometropic group, and superior RNFL quadrant in the myopic anisometropic group were significantly thinner in amblyopic eyes compared to their fellow eyes (P < .05). RNFL thickness did not differ between strabismic amblyopic, anisometropic amblyopic, and control eyes (P > .05). In the anisometropic group, a significant correlation was found between mean RNFL thickness and refraction (P < .05). Conclusion: Amblyopia is not associated with a decrease in RNFL thickness in strabismic or anisometropic amblyopia. In the anisometropic group, the inter-eye diff erences in RNFL thickness parameters seemed to be related to the refraction diff erences between the amblyopic eyes and their fellow eyes.