Purpose: To analyze the macular ganglion cell-inner plexiform and retinal nerve fiber layer thicknesses after vitrectomy with the inverted flap technique for idiopathic macular holes. Methods: A prospective study was conducted on 28 eyes treated with surgery for idiopathic macular holes. The inverted internal limiting membrane flap technique assisted with Brilliant Blue staining (0.05%) was performed. Ophthalmologic examinations and quantitative analysis of the macular ganglion cell complex thickness were performed at baseline,1 and 3 months after surgery. Results: The preoperative mean thicknesses of the ganglion cell-inner plexiform layer and ganglion cell-inner plexiform layer + retinal nerve fiber layer were 88.9 and 124.8 μm, respectively. The mean ganglion cell-inner plexiform layer thicknesses at 1 and 3 months after surgery were reduced to 72.8 and 65.2 μm, respectively (p<0.001 and p<0.001, respectively). The mean postoperative ganglion cell-inner plexiform layer + retinal nerve fiber layer thickness was also reduced at 1 and 3 months (108.8 and 99.3 μm, respectively; p<0.001 and p<0.001, respectively). No significant difference was found between the preoperative and postoperative best-corrected visual acuities at 1 and 3 months (p<0.73 and p<0.14, respectively). Conclusion: The macular ganglion cell-inner plexiform layer and ganglion cell-inner plexiform layer + retinal nerve fiber layer thicknesses were significantly reduced after vitrectomy with the inverted flap technique assisted with Brilliant Blue staining (0.05%) for idiopathic macular holes.