<p>Cyclophosphamide is a widely used chemotherapeutic agent known for its effectiveness in treating various cancers; however, it is associated with significant immunosuppressive side effects. This study investigates the potential of sakuranetin, a natural flavonoid, in mice under cyclophosphamide-induced immunosuppressive conditions. Mice were grouped into four groups: one control group, two treated with cyclophosphamide and two sakuranetin-treated groups receiving different doses (10 and 20 mg/kg). Immune organ indices, lymphocyte proliferation, hematological parameters, nitric oxide levels, cytokines (tumor necrosis factor alpha-TNF-α, interferon γ-IFN-γ), interleukins (interleukin-1β-IL-1β, IL-4, IL-6), immunoglobulin G (IgG), IgM levels, plague-forming cells quantification, qualitative hemolysis, and delayed-type hypersensitivity were assessed. Cyclophosphamide significantly (<i>P</i> &lt; 0.05) reduced immune organ indices, lymphocyte proliferation, changes in hematological parameters, and nitric oxide levels. Treatment with both sakuranetin doses restored these parameters and normalized cytokine, IgG, and IgM levels (<i>P</i> &lt; 0.05). Sakuranetin significantly (<i>P</i> &lt; 0.05) improved the immunomodulatory action with elevated immune response with downregulation in immune response mediated by cells. Sakuranetin effectively counteracts cyclophosphamide-induced immunosuppression by modulating the IFN-γ, TNF-α, IgG, and interleukin pathway, suggesting its potential as a protective agent.</p> Graphical Abstract <p></p>