Blood outgrowth endothelial cells (BOECs) are important tools when investigating diagnostic and therapeutic approaches for vascular disease. In this protocol, mononuclear cells are isolated from peripheral blood and plated on type I collagen at ĝ̂1/4135,000 cells per cm 2 in endothelial cell differentiation medium. On average, 0.34 colonies of endothelial cells per milliliter of blood can be obtained. Colonies of endothelial cells become visible after 14g-28 d. Upon confluence, these rapidly expanding colonies can be passaged and have been shown to propagate up to 10 18 -fold. Isolated BOECs are phenotypically similar to vascular endothelial cells, as revealed by their cobblestone morphology, the presence of endothelial cellg-specific Weibel-Palade bodies and the expression of endothelial cell markers such as VE-cadherin. The protocol presented here also provides a particularly useful tool for the ex vivo assessment of endothelial cell function from patients with different vascular abnormalities.