Objective To isolate ES cell colonies on a feeder layer. Methods Mouse embryos of 13-14 days were for the cultivation of primary mouse embryo fibroblasts (PMEF) which turned to be a feeder layer for blastocysts. Results The inner cell mass (ICM) of blastocysts grew well on the feeder layer of PMEF.The proliferation rate of ICM was 57%. After 48 hours of ICM disagregation ES colonies began to appear,showing strong AKP activity. Conclusion It is a preliminary success for the feeder layer to produce embryonic stem cell lines.