Endothelial progenitor cells (EPCs) can be isolated from adult peripheral and umbilical cord blood and expanded exponentially ex vivo. In contrast, human umbilical vein endothelial cells (HUVECs) or human aortic endothelial cells (HAECs) derived from vessel walls are widely considered to be differentiated, mature endothelial cells (ECs). However, similar to adult- and cord blood-derived EPCs, HUVECs and HAECs derived from vessel walls can be passaged for at least 40 population doublings in vitro. Based on this paradox, we tested whether EPCs reside in HUVECs or HAECs utilizing a novel single cell deposition assay that discriminates EPCs based on their proliferative and clonogenic potential. We demonstrate that a complete hierarchy of EPCs can be identified in HUVECs and HAECs derived from vessel walls and discriminated by their clonogenic and proliferative potential. This study provides evidence that a diversity of EPCs exists in human vessels and provides a conceptual framework for determining both the origin and function of EPCs in maintaining vessel integrity.