Objective To isolate and identify the rat amniotic cells and to explore their stem cell characteristics, so as to find a new cell source for stem cell transplantation. Methods Rat amnion tissue was mechanically separated from 18-18.5-day old SPF pregnant rats and 0.25% trysin-EDTA digestion was used to obtain amniotic cells. The isolated cells were cultured with DMEM D-glucose added with 10 μg/L EGF. Flow cytometry was used to detect the mesenchyme stem cellsurface markers. Neural stem cell surface markers of the rat amniotic cells were detected using immunofluorescence staining. Results The rat amniotic membrane tissue was separated and mesenchymal stem cells were cultured successfully. The cells were ameboid-shaped and showed adherent growth, and can stably proliferate in short term. After culture, the cells expressed stem cell markers e.g. Oct-4 and Sox-2, mesenchymal stem cell markers e.g. vimentin, embryonic stem cell markers e.g. SSEA-4, neural stem cell marker e.g. nestin, and could also express neurotrophic factors, such as BDNF and NGF. Conclusions Rat amniotic cells express mesenchymal stem cells markers, neural stem cell markers and neurotrophic factors, therefore, provide an experiment basis for further research and application of rat amniotic cells in stem cell transplantation.