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Background: We investigated the effect of human serum albumin (HSA) on human umbilical cord blood (UCB) CD34 + hematopoietic stem/progenitor cells (HSPCs) cultured in vitro and transplanted in vivo ., Methods: Human umbilical cord blood mononuclear cells were obtained by density gradient centrifugation. CD34 + cells were then sorted by CD34 conjugated magnetic microbeads. The sorted cells were cultured with or without HSA for 8 days in vitro . After 8 days, all cells were harvested for flow phenotyping and colony formation cell (CFC) experiments. The cells were injected into immunodeficient mice (NOD/Shi-scid/IL2Rγnull, NOG) via intravenous injections. From 4 weeks post-transplantation, flow cytometry was used to calculate human cell chimerism in the peripheral blood (PB) every 2 weeks. Flow phenotyping of human cell chimerism in bone marrow and spleen was calculated 16 weeks post-transplantation., Results: Compared to the control group, CD34 + cells cultured with HSA increased significantly in vitro . The long-term engraftment of HSPCs and the hematopoietic multilineage reconstruction capacity were preserved by HSA. Normal engraftment of human cells could be maintained via HSA treatment could maintain normal engraftment of human cells in recipient PB., Conclusions: Here, we found that HSA was beneficial to maintaining CD34 + cell expansion and short-term colony formation in vitro and optimizing multilineage reconstitution in immunodeficient mice in vivo ., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://atm.amegroups.com/article/view/10.21037/atm-22-6383/coif). The authors have no conflicts of interest to declare., (2023 Annals of Translational Medicine. All rights reserved.)