The aim of the current study is to evaluate retinal nerve fiber layer (RNFL) thickness, macular parameters and ganglion cell thickness (GCL+) in unilateral anisometropic amblyopic eye in relation to the normal healthy fellow eye and control one using spectral domain optical coherence tomography (SD-OCT). This study included 56 eyes of 28 cases aged from 8-30 years old subdivided into: 20 amblyopic eyes of patients with unilateral anisometropic amblyopia (group A), 20 sound fellow eyes of the unilateral anisometropic amblyopic patients (group B) and 16 emmetropic eyes of normal non amblyopic control (group C). Anisometropic amblyopic patients further subdivided according to refractive error into hypermetropic, myopic, and astigmatic amblyopic subgroups. All cases underwent detailed ophthalmic examination including visual acuity by Snellen's chart, cycloplegic refraction, ocular motility and alignment test, slit lamb biomicroscopy, intraocular pressure, fundus examination and OCT imaging by SD-OCT. Regarding peripapillary RNFL thickness, macular parameters including (central macular thickness (CMT), average macular thickness and macular volume), and GCL+ thickness, there was no significant difference among the amblyopic, fellow eyes, and normal eyes of non-amblyopic group, P˃0.05 . By further analysis to compare subgroups according to the refractive error between amblyopic and their fellow eyes, peripapillary RNFL thickness revealed a significant increase in amblyopic eyes of hypermetropic (mean 121.750±8.647μm) and astigmatic subgroups (mean 111.888±7.043μm) compared with their fellow eyes (116.125±8.509 and 107.333±6.480) respectively, but it was significantly thinner in myopic amblyopic eyes (mean 81.333±23.352μm) than that of their fellow eyes (mean 110.00±17.578). CMT revealed a significant increase in myopic amblyopic eyes (mean 199.00±26.851μm) compared with their fellow eyes (mean 191.00±26.00μm). Other OCT parameters did not significantly differ between amblyopic and their fellow eyes in different subgroups. OCT parameters including peripapillary RNFL thickness, macular parameters, and GCL+ thickness showed no significant difference among the amblyopic, fellow eyes, and normal eyes of non-amblyopic group. Hypermetropic and astigmatism amblyopic eyes showed increased peripapillary RNFL thickness than their fellow eyes, while myopic amblyopic eyes had decreased peripapillary RNFL thickness than their fellow eyes. CMT revealed a significant increase in amblyopic eyes of the myopic anisometropic subgroup compared with their fellow eyes.