无饲养层培养人胚胎干细胞方法的建立

人胚胎干细胞(human embryonic stem cell，hES细胞)是当前医学研究的热点之一．然而hES细胞培养条件苛刻，通常需要采用鼠胚胎成纤维细胞(mouse embryonic fibroblast，MEFs)饲养层来维持其未分化状态，成为目前hES细胞研究的瓶颈之一、本实验成功地将hES细胞接种在细胞外基质包被的六孔板上培养，传代20次后细胞仍然保持良好的未分化状态，各种hES细胞生物学特性(如表面标志物SSEA-3、SSEA-4、TRA-1-60和TRA-1-8l，OCT-4，碱性磷酸酶及体内外分化潜能等)均无改变；其冻存、复苏效果与生长在饲养层上的hES细胞无明显差异．因此，该无饲养层培养体系可以用于培养hES细胞，并为hES细胞转基因研究及大规模培养打下良好的基础．

Establishment of a Feeder-free System for Human Embryonic Stem Cells

Human embryonic stem cell (hES) research is one of hot spots in cur re nt medical science.However,culturing hES cells requires critical conditions and mouse embryonic fibroblast (MEF) feeders are commonly used to maintain the uniqu e characteristics of hES cells,which becomes one of the bottlenecks in hES cell research.In this experiment,hES cells cultured on MEF feeders were successfully transferred to Matrigel coated six well plates and still maintained undifferen tiated for at least 20 population doublings.The biological characteristics of un differentiated hES cells (e.g.expression of OCT 4,alkaline phosphatase(AKP),and surface markers including SSEA 3,SSEA 4,TRA 1 60,TRA 1 81,in vitro and in vivo differentiation potency) were unchanged.The efficiencies of freezing and t hawing hES cells on Matrigel were comparable to those on feeders.Therefore,the f eeder free system is suitable for hES cells,which lays a good foundation for hE S cells transfection and scale up production.