Background Interest in the biology of endogenous progenitor cells (EPCs) continues to grow as evidence of their role in vascular repair mounts. EPC enumeration requires specialized laboratory techniques and is performed immediately after sample acquisition, limiting the clinical contexts in which EPC enumeration can be performed and the ability to increase sample sizes through multi-center participation. Methods We compared the numbers of EPCs enumerated in samples processed immediately after acquisition (n = 36) with EPCs enumerated in specimens stored for 24 hours or after cryopreservation of mononuclear cells (MNC) using two EPC identification strategies: cell surface marker expression (CD133/CD34) and aldehyde dehydrogenase activity (ALDHbr cells). Results EPCs assessed in fresh samples correlated with EPCs enumerated after whole blood storage (r = 0.699 for CD133+CD34+ cells, r = 0.880 for ALDHbr cells, P < 0.005 and P < 0.0001, respectively) or mononuclear cryopreservation (r = 0.590 for CD133+CD34+ cells, r = 0.894 for ALDHbr cells, P < 0.0001 for each); however, correlation based on assessment of ALDHbr cells was higher (P < 0.0003 for comparison of correlation coefficients). Initial results from a multi-site clinical trial suggest that EPC enumeration after mononuclear cell cryopreservation is feasible. Conclusion EPC analysis based on ALDH activity is reproducible, even after extended whole blood storage or MNC cryopreservation.