Objective To improve methods of culturing rat myocardial microvascular endothelial cells (RMMVECs) EM>in vitro/EM> and provide experimental data for studying the structures and functions of RMMVECs. Methods SD rats of 10_15 days old were selested. Explants cultivation of cardiac tissues was used to obtain the primary cells. In the subculturing, the cell cultures were purified by different speeds of attachment and digestion from contaminant cells combined with morphological observation. The purified cells were identified as endothelial cells by morphology and by positive immunocytochemistry for factor Ⅷrelated antigen, a marker for endothelial cells. Results The RMMVECs were cultured and purified successfully which grew in a polygonal or cobblestone pattern and were positive for the immunocytochemical staining of factor Ⅷrelated antigen. Conclusion The methods of culturing RMMVECs were improved, RMMVECs with high purity were o