The study was performed to evaluate the suitability of glycerol-cryopreserved human amniotic membrane (HAM) as a surgical patch, far from its common use in ophthalmic surgery. In vivo experiments in rat models were performed to study the degradation patterns, biocompatibility, postoperative tissue formation and its suitability for abdominal wall closure. Degradation and thickness of the membranes were assessed over a period of 60 days after subdermal implantation of monolayer and multilayer HAM in 96 immunocompetent and immunosuppressed rats. The tissue response was mild, and histological analysis evaluated that multilayer application and immunosuppression prolonged graft survival significantly. In a second rat model, another 18 animals were monitored over a period of 28 days after abdominal wall reconstruction with multilayered HAM. Polypropylene mesh (Prolene) and polyglactin910/polydioxanon patches (Ethisorb) served as controls. Gross examination and histological analysis proved that multilayer HAM was a sufficient material for abdominal wall closure in comparison with the polypropylene mesh and was superior to the polyglactin910/polydioxanon patch. Additionally, significantly reduced postoperative intraabdominal adhesions were observed when compared to the polyglactin910/polydioxanon patch. This study demonstrates that HAM is a biocompatible, resorbable surgical patch in a rat xenotransplantation model and serves as a mechanically sufficient material for abdominal wall closure in a small animal model. These findings are encouraging and justify further research for the use of cryopreserved human amniotic membrane in soft tissue repair.