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Objective: To optimize the fermentation process of ginseng and wolfberry fermentation liquid and study its regulating effect on the immune function of mice. Methods: The fermentation process was optimized by single factor test and Box-Behnken test with total acid content as response value and fermentation time, fermentation temperature, strain proportion and inoculum amount as independent variables. The mice were induced with 80 mg/(kg·d) cyclophosphamide to establish an immunocompromised mouse model. The mice were treated with low (5.0 mL/kg), medium (10.0 mL/kg) and high (15.0 mL/kg) doses of experimental fermentation liquid solution, respectively. The contents of immunoglobulin G (IgG), interleukin-4 (IL-4), interleukin-10 (IL-10) and interferon-γ (interferon-γ) in serum were determined. The relative expression levels of IFN-γ and tumor necrosis factor-α (TNF-α) were measured. The phagocytosis ability of mouse phagocytes and its effect on organ index were detected by carbon clearance method. Results: The fermentation process was optimized as follows: Fermentation time 72 h, fermentation temperature 37 ℃, inoculation amount 5%, strain ratio 5:4.5:0.5, total acid content 1.23 g/100 g. The levels of IFA-γ, TNF-α, IL-4 and IgG in the serum of immunosuppressed mice were significantly up-regulated by the fermentation liquid, and the levels of IL-10 were down-regulated. The mass concentration of IL-10 and IFN-γ in the serum of mice with high dose could be restored to 72.26 and 213.47 pg/mL, respectively. The mass concentration of TNF-α and IgG reached 246.13 pg/mL and 119.89 ng/mL, which were close to or higher than those of solvent control group, and significantly enhanced the phagocytic ability of phagocytic cells. Conclusion: The ginseng and wolfberry fermented liquid had a positive regulatory effect on the immune function of mice.