Interleukin 12 (IL12), a heterodimer cytokine composed of IL12A (p35) and IL12B (p40), is mainly produced by dendritic cells and macrophages in response to antigenic stimulation and its receptor is a protein complex encoded by IL12Rβ1 and IL12Rβ2. IL12 signaling regulates innate and adaptive immunity, influencing the proliferation and activation of cytotoxic lymphocytes and natural killer (NK) cells, stimulating the production of interferon (IFN)-γ, and promoting the differentiation and production of Th1-associated immunoglobulins. IL12 is a promising anti-tumor drug capable of transforming the tumor microenvironment from “cold” to “hot”. To accelerate discovery in this space, Biocytogen has independently developed IL12RB1 and IL12RB2 single KI mice and mated them to subsequently B-hIL12RB1/hIL12RB2 double KI mice to develop a fully humanized model for testing IL12-analog drugs. First, the mRNA expression levels of human IL12RB1 and human IL12RB2 in B-hIL12RB1/hIL12RB2 mice were quantified using RT-qPCR. Additionally, immunophenotyping by flow cytometry shows that the humanization of IL12RB1 and IL12RB2 did not affect the proportions and distribution of immune cells. To validate the functionality of the humanized IL12Rβ1 and IL12Rβ2 receptor complex, CD4⁺ T cells sorted from splenocytes of C57BL/6 and homozygous B-hIL12RB1/hIL12RB2 mice were stimulated with human IL12 in vitro, and IFN-γ production was detected by ELISA. The results indicated that human IL12 induced a significant increase in IFN-γ levels in the B-hIL12RB1/hIL12RB2 mice, while no such effect was observed in wild-type mice. Furthermore, to evaluate the in-vivo toxicity of human IL12 analogs, B-hIL12RB1/hIL12RB2 mice were treated with human IL12 at different doses and for varying durations. Body weight was measured daily during treatment. At the end of the experiment, the weights of the liver and spleen were measured, and pathological changes in the liver and spleen were assessed through HE staining. Meanwhile, serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the mice were detected using ELISA. In the in vivo toxicity study, human IL-12 administration resulted in a progressive decline in body weight in treated mice over time. Additionally, human IL12 treatment was associated with an increase in the ratios of liver weight to body weight and spleen weight to body weight. Pathological changes in the liver and spleen were also observed, alongside an upward trend in ALT and AST activity. These results suggest the successful humanization of IL12RB1 and IL12RB2 signaling in B-hIL12RB1/hIL12RB2 mice, enabling evaluation of the preclinical toxicity of human IL12 analogs.