Objective To optimize the culture condition for porcine bone marrow-derived endothelial progenitor cells (EPCs), so as to lay a foundation for future study. Methods Bone marrow was collected from porcine ilium (n=5). Mononuclear cells (MNCs) were separated by density centrifugation and were induced to differentiate into EPCs in vitro. The influences of different cell inoculation densities (2×103/cm2, 5×103/cm2, 1×104/cm2, and 2×104/cm2), basic medium (EGM, medium 199, and DMEM), FBS (5%, 10%, 20%, and 30%), and combinations of cytokines (vascular endothelial growth factor \[VEGF\]+basic fibroblast growth factor \[bFGF\], VEGF+stromal-derived factor \[SDF\], VEGF+bFGF+SDF, VEGF+bFGF+insulin-like growth factor \[IGF\]+epidermal growth factor \[EGF\], and VEGF+bFGF+SDF+IGF) on the proliferation and migration of EPCs were evaluated. EPCs were identified by morphology observation, fluorescent staining, and immunohistochemical method. Results EPCs with the highest proliferation and migration ability were obtained under following condition: at a density of 1×104/cm2 in M199 medium supplemented with 10% FBS and VEGF+bFGF+SDF+IGF. Furthermore, the percentage of double positive cells stained by Dil-ac-LDL and FITC-UEA-1 was higher than 76%, and these cells were also positively stained for CD133, CD34 and KDR immunohistochemically. Conclusion Optimization of in vitro culture condition of porcine EPCs can increase the cell amount and improve their functions, paving a way for future related studies.