Objective To observe the changes in hematopoietic and immune function at 6 months post 4 Gy or 6 Gy γ-ray total body irradiation (TBI) in C57BL/6 J mice. Methods C57BL/6 J mice of 8- 12 weeks of age were irradiated with 4 Gy or 6 Gy TBI. At 6 months after irradiation exposure, the peripheral blood cell count and classification, immune organ coefficient, immune phenotyping of the peripheral blood and spleen cells, and expression level of splenic T cell p16 were detected. Results Six months after irradiation exposure, the peripheral blood leukocyte and red blood cell count in the mice exposed to TBI remained significantly lower than those in the control group. There was no significant difference in the count of platelets in the irradiated mice compared to the control group. The result of the peripheral blood cell classification showed that the proportion of neutrophils in the irradiated mice group increased and the proportion of lymphocytes decreased, indicating a skew in blood cell differentiation. Compared with the control group, the thymus coefficient in the irradiated mice was significantly decreased at 6months post 6 Gy TBI ( P <0. 05). There was no significant difference in the spleen coefficient. Phenotyping of the peripheral blood cells was performed by flow cytometry. The result showed that the proportion of B lymphocytes and NK cells in the irradiated mice remained lower than those in the control group, while the proportion of CD4 lymphocytes in the irradiated group was higher than that in the control group. There was no significant difference between CD8 lymphocytes and monocyte levels in the irradiated group compared with the control group. The proportion of B220 cells was clearly decreased in the irradiated mice compared with the control group based on the splenic immune phenotyping assay ( P <0. 01). Compared with the control group, the expression of p16 in the splenic T cells of the 6 Gy TBI mice was increased but not significantly ( P > 0. 05). Conclusions Hematopoietic immune phenotyping indicated the recovery at 6 months after the sublethal dose of TBI but the neutrophils in the peripheral blood continued to be increased, and the thymus coefficient and the proportion of B lymphocytes in the peripheral blood and spleen were still lower than those in the control group, suggesting a long-term injury. These experimental result provide basic data for the study of total body irradiation-induced long-term injury of hematopoietic immune function.