Abstract: Abstract Objective：Explore an economic and reliable, stable, high purity of endothelial progenitor cells in culture methods.Methods：Rat bone marrow-derived EPCs were separated by density gradient centrifugation and cultured，then plated on 25ml culture flask adherent culture with human fibronection, 48h no adherent cells were collected cultured for 7 days, and then identified by being observed the experirment of uptaking DiL-acLDL and combining FITC-UEA-1 by fluorescence microscope . Also Flow Cytometry was performed to analyze the expression of CD133、flk-1、VE-cadherin(CD144) and CD31 dynamically.Results：48h no adherent cells cultured,the cells were small and spindle or polygonal in shape.Large numbers of typical endothelial progenitor cells coloy-forming unites were found. Cells DiI-acLDL uptake and FITC-UEA-1 combining fluorescent staining positive rate（87.0±6.0）%.Flow cytometry to detect cell surface antigen CD133, flk-1, VE-cadherin and CD31 positive rates were 61.01%, 8.55%,8.20%,6.18%. Conclusions：Differential attachment method is a reliable, stable, high purity of endothelial progenitor cells culture methods. Provide methodological references for further research.

Key words: Differential attachment method, Rat, Mononuclear cells, Endothelial progenitor cells