• The recombinant-activating gene 2 (Rag2) plays an important role in the rearrangement and recombination of the genes of immunoglobulin and T cell receptor molecules during the initiation of V(D)J recombination. Loss of Rag2 protein leads to no mature T and B cells, the critical components of the adaptive immune system.
• Il2rg is known as the interleukin receptor common gamma chain, which is an important signaling component of many interleukin receptors, including IL2, IL4, IL7, IL9, IL15 and IL21. IL15 is the main cytokine promoting NK cell differentiation and maturation. Knockout of the Il2rg gene will result in no mature NK cells in mice.
• Double knockout of Rag2 and Il2rg genes in mice results in an absence of mature T/B/NK cells, thereby creating a severe immunodeficiency mouse model. It can be used to support research in many areas including immune system defections, virology research, inflammation research, cancer research, xenograft/transplant host etc.

The size and weight of thymus and spleen of B-CDG mice were significantly reduced compared to that of wild-type C57BL/6 mice. Thymuses and spleens were isolated from wild-type C57BL/6 mice and B-CDG mice (n=3, 7-week-old). A. Gross anatomy of thymuses; B. The size of spleen and thymus; C. Comparison of organ coefficients of spleen and thymus in wild-type C57BL/6 mice and B-CDG mice. Results showed that thymuses were not visualized in B-CDG mice. The size and weight of spleens in B-CDG mice were significantly reduced compared to that in wild-type C57BL/6 mice.  

Lymph nodes were not visualized in B-CDG mice dyed by Evans blue. Wild-type C57BL/6 mice and B-CDG mice (n=3) were sacrificed and intravenously injected with 10% Evens blue in the planta pedis. The red arrows indicate the lymph nodes in inguinal, axillary and mesenteric site of the mice. No lymph node was visualized in B-CDG mice.

Frequency of leukocyte subpopulations in bone marrow by flow cytometry. Bone marrow cells were isolated from male wild-type C57BL/6 mice and homozygous B-CDG mice (n=3, 6-week-old). A. Flow cytometry analysis of the bone marrow cells was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. T cells, including CD4+ T cells, CD8+ T cells and Tregs, B cells and NK cells were not detectable in B-CDG mice. Frequency of neutrophils, monocytes and macrophages were increased in B-CDG mice compared to that in C57BL/6 mice, demonstrating that the development of T cells, B cells and NK cells in bone marrow were completely inhibited in B-CDG mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.001.

Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from male wild-type C57BL/6 mice and homozygous B-CDG mice (n=3, 6-week-old). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. T cells, including CD4+ T cells, CD8+ T cells and Tregs, B cells and NK cells were not detectable in B-CDG mice. Frequency of dendritic cells, neutrophils, monocytes and macrophages were increased in B-CDG mice compared to that in C57BL/6 mice, demonstrating that the development of T cells, B cells and NK cells in spleen were completely inhibited in B-CDG mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.001.

Frequency of leukocyte subpopulations in blood by flow cytometry. Blood cells were isolated from male wild-type C57BL/6 mice and homozygous B-CDG mice (n=3, 6-week-old). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. T cells, including CD4+ T cells, CD8+ T cells and Tregs, B cells and NK cells were not detectable in B-CDG mice. Frequency of dendritic cells, neutrophils and macrophages were increased in B-CDG mice compared to that in C57BL/6 mice, demonstrating that the development of T cells, B cells and NK cells in blood were completely inhibited in B-CDG mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.001.

Comparison of the irradiation tolerance of B-CDG mice and B-NDG mice by using different doses of X-rays. B-NDG mice and B-CDG mice were exposed to graded X-ray doses; survival and body weight were monitored up to 30 days post-irradiation. (A-B) Survival rate and body weight change of B-NDG mice. (C-D) Survival rate and body weight change of B-CDG mice. When the irradiation dose reached 2.5 Gy or more, all of B-NDG mice died 2 days after irradiation. Only 1/6 of the B-NDG mice could survive until 30 days after irradiation at a dose of 1 Gy. All B-CDG mice survived until 30 days when the irradiation dose was 2.5 Gy; some of them could survive until 30 days when the irradiation dose was 4 Gy; and all B-CDG mice died 9 days after irradiation when the dose was 7 Gy. Across comparable doses, B-CDG mice demonstrate substantially greater whole-body irradiation tolerance than B-NDG mice. Values are expressed as mean ± SEM.

Complete Blood Count (CBC) was measured at day 7 and day 30 post-irradiation. Blood were collected on the indicated days and tested by CBC. Across groups, WBC and differential metrics showed post-irradiation suppression with partial recovery by day 30. B-CDG mice had not recovered by day 30 and exhibited slower WBC recovery. Values are expressed as mean ± SEM.

RBC indices and erythropoiesis markers were quantified at day 7 and day 30 post-irradiation. Blood were collected on the indicated days and tested by CBC. The results showed that there was a clear dose-dependent impact on RBC parameters, with high-dose groups showing reduced RBC and erythropoietic capacity at day 7, followed by substantial recovery by day 30 (high-dose groups recover more slowly). Irradiation induced transient anemia/erythropoietic suppression that largely normalized by day 30, with slower normalization at higher doses. Values are expressed as mean ± SEM.

Platelet parameters were measured at day 7 and day 30 post-irradiation. Blood were collected on the indicated days and tested by CBC. The results showed that higher dose of irradiation caused greater platelet impact, with marked reduction in platelet number/platelet production capacity at day 7 in higher-dose groups and near-recovery by day 30, again with slower recovery at higher doses. Values are expressed as mean ± SEM.

Several organs were harvested and measured by H&E staining (400×) at day 30 post-irradiation. All groups showed significant hematopoietic stress and compensatory responses (e.g., extramedullary hematopoiesis; marrow cellularity changes). The ovarian atrophy and inflammatory infiltrates suggested gonadal radiosensitivity even when animals survive. Red arrows: extramedullary hematopoiesis; Green arrows: hyperpigmentation; Bold black arrows: inflammatory infiltrates; Thin black arrows: ovarian atrophy and proliferation of stromal cells; Orange arrows: Increased bone marrow cellularity with an elevated myeloid-to-erythroid (M:E) ratio; Black triangle: hematoma.

Tumor formation of SHP-77 human ductal breast cancer cell line in B-CDG mice. SHP-77 cells (2×10⁶) were subcutaneously inoculated into B-CDG mice (female, 7-week-old, n=6). Results showed that SHP-77 cells could successfully form tumors in B-CDG mice. Values are expressed as mean ± SEM.

Tumor formation of BT-474 human ductal breast cancer cell line in B-CDG mice. BT-474 cells (1×10⁷) were subcutaneously inoculated into B-CDG mice (female, 7-week-old, n=6). Results showed that BT-474 cells could successfully form tumors in B-CDG mice. Values are expressed as mean ± SEM.

Tumor formation of AsPC-1 human pancreatic cancer cell line in B-CDG mice. AsPC-1 cells (2×10⁶) were subcutaneously inoculated into B-CDG mice (female, 7-week-old, n=6). Results showed that AsPC-1 cells could successfully form tumors in B-CDG mice. Values are expressed as mean ± SEM.

Tumor formation of LNCap Clone FGC human prostate cancer cell line in B-CDG mice. LNCap Clone FGC cells (2×10⁶) were subcutaneously inoculated into B-CDG mice (male, 7-week-old, n=6). Results showed that LNCap Clone FGC cells could successfully form tumors in B-CDG mice. Values are expressed as mean ± SEM.

Tumor formation of 22Rv1 human prostate cancer cell line in B-CDG mice. 22Rv1 cells (1×10⁶) were subcutaneously inoculated into B-CDG mice (male, 7-week-old, n=6). Results showed that 22Rv1 cells could successfully form tumors in B-CDG mice. Values are expressed as mean ± SEM.