Platelet-rich plasma (PRP) containing autologous growth factors is applied in regenerative medicine, but the lack of an optimized PRP preparation protocol causes unstable therapeutic effects. The aim of this study was to optimize the PRP preparation method and compare the effects of PRP from different preparation methods in restoration of erectile function in a rat model. The in vivo experiments used Sprague–Dawley male rats (n = 24), which were randomly divided into four groups of equal numbers: group I underwent sham operation, while the remaining three groups underwent bilateral CN crush. Crush injury groups were treated at the time of injury with an application of general PRP, optimized PRP [with the largest amount of platelet-derived growth factor (PDGF)–AB] or normal saline-only injection in the corpus cavernosum, respectively. Four weeks later, erectile function was assessed by CN electrosimulation, and penile tissue was collected for histology. Results demonstrated that in the PRP group prepared with the ACD-A anticoagulant, chitosan and incubated at −20°C for 15 days had the largest amount of PDGF-AB and showed a synergistic effect on release (p < 0.05). Functional outcome measurement and immunofluorescence staining for the dorsal nerve revealed that improvement after bilateral CN injury occurred in the optimized PRP group (p < 0.05). It was concluded that optimized PRP with a high level of growth factors was more stable, and its injection into the corpus cavernosum facilitated recovery of erectile function. Copyright © 2013 John Wiley & Sons, Ltd.