The possibility of maintaining, manipulating, and expanding human hematopoietic stem cells in ex vivo culture could help to provide patients with autologous and allogeneic stem cell transplants improved in purity and performance and could offer access to gene therapy of the hematopoietic system. Recent advances in the ex vivo culture of immature human hematopoietic progenitor cells and human hematopoietic stem cells have led to experimental evidence for the qualitative and quantitative maintenance and possible numerical expansion of hematopoietic stem cells in ex vivo culture, making ex vivo graft engineering a realistic possibility. This review summarizes recent developments in the field, their regulatory implications and their application in hematopoietic gene therapy.