It has been found that post-radiation mouse embryonic fibroblasts can well maintain the pluripotency in human embryonic stem cells. However, the molecular mechanism remains unclear. In the present study, the new protein expression profile of post-radiation mouse embryonic fibroblasts was analyzed by immobilized pH gradient 2-dimensional polyacrylamide gel electrophoresis. Image analysis following silver staining revealed (969 57) vs. (1085 107) spots from post-radiation mouse embryonic fibroblasts and pre-radiation ones, respectively. Some newly expressed proteins, which were only abundantly present after irradiation, were subjected to peptide mass fingerprint analysis and identified using MALDI-TOF-MS, SWISS-PROT database, and RT-PCR. Several of those proteins were preliminarily identified to participate in cytokine secretion, cell signal transduction, transcriptional regulation, and apoptosis, etc., which suggested that inactive post-radiation mouse embryonic fibroblasts expressed some new proteins that may underlie the molecular mechanisms to maintain the pluripotency in human embryonic stem cells. (C) 2004 Elsevier Inc. All rights reserved.