Aim: To induce embryonic stem (ES) cells into nerve cells in vitro by 5-step method. Methods: Firstly, ES cells were inoculated into culture bottles pretreated with gelatin, and cultured for 3 days without enriched cells, followed by suspension culture for 4 days. After the cells were transformed into embryoid body (EB), the EB was cultured in the non-serum medium for 6 days to enrich nestin-positive cells. The cells were amplified by cultured in basic fibroblast growth factor for 6 days. Finally, the cells were induced and differentiated by nicotinamide for 6 days. Results: Among the cells that had participated in the entire induction procedure, the number of immature neurons that expressed Tui-1 was up to 80% - 90%, the number of astrocytes expressing glial fibrillary acidic protein up to 5% - 10%, and the number of mature neurons expressing microtubule associated protein-II (MAP-II) up to 5% - 10%. Conclusion: Five-step method makes the differentiated procedure of EB cells divided into different stages, a few astrocytes and mature neurons, which may be useful for further neurobiology study as well as potential transplantation therapy.