PURPOSE: To report a case of long-term postoperative vitreous hemorrhage (PVH) of proliferative diabetic retinopathy after pars plana vitrectomy (PPV), which is most likely to be associated with neovascularization (NV) formed at the incision site of vitrectomy, and to recommend a better surgical technique for the diagnosis and treatment of such patients. METHODS: Intraoperative fundus fluorescein angiography (FA) was performed using a customized intraoperative blue light excitation source and observed by a three-dimensional (3D) surgical observation system to explore the causes of recurrent PVH. RESULTS: Intraoperative 3D-FA clearly showed the complete fluorescence image of the inward growing scleral NV at the PPV incision site and guided the surgical treatment of intraocular retinal laser photocoagulation of the NV. CONCLUSION: Intraoperative 3D-FA has a certain role in the diagnosis and treatment of unexplained PVH after PPV, and is a promising surgical auxiliary technology.