Background: Immune responses inhibit invasion by pathogens and antigens. Thus, it is important to promote the immune response in immunosuppressed patients. Objective: To examine whether electroacupuncture (EA) promotes the immune response by regulating the downstream regulatory element antagonist modulator / nuclear factor kappa B (DREAM/NF-κB) signalling pathway in a mouse model of cyclophosphamide (CP)-induced immunosuppression, and determine the most effective frequency. Methods: Twenty-four Kunming mice were intraperitoneally injected with CP to establish an immunosuppression model and six mice were injected with the same volume of normal saline as a control. The 24 mice were randomly divided into four groups: manual acupuncture, 2 Hz EA treatment, 100 Hz EA treatment and alternating 2/100 Hz EA treatment. After EA treatment for 3 days, immune response, natural killer (NK) cell toxicity and the expression of cytokines and DREAM/NF-κB were assessed. Results: EA treatment, especially at alternating 2/100 Hz frequency, improved spleen and thymus indices, increased lactate dehydrogenase and acid phosphatase levels, promoted concanavalin A- and lipopolysaccharide-induced splenocyte proliferation, increased NK cell toxicity and ameliorated CP-induced immunosuppression in mice. Additionally, 2/100 Hz EA treatment increased interleukin (IL)−2, IL-6, IL-12, tumour necrosis factor-α and interferon-γ levels and decreased IL-10 levels in CP-induced immunosuppressed mice. Finally, it was found that 2/100 Hz EA treatment increased p-IκBα and NF-κB expression and decreased DREAM and IκBα expression, suggesting that this treatment activates the NF-κB signalling pathway. Conclusion: 2/100 Hz EA treatment might be an effective way to enhance immune responses in CP-induced immunosuppressed mice.