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Radiofrequency (RF) can destroy local tumor tissue with less damage to the intervening tissue. The effect of RF on immune function of tumor-bearing body needs to be further investigated. This study was designed to evaluate the impact of RF on splenocyte immunity of mice bearing H22 liver cancer through detecting splenocyte proliferation, cytotoxicity, and expression pattern of cytokines T helper type 1 (Th1)/Th2.A total of 24 BALB/c mice were randomized into 4 groups: RF, surgical resection, tumor-bearing control, and normal control groups. The proliferation of splenocytes was detected by MTT assay, and the cytotoxicity of splenocytes was assayed by double-color flow cytometry (FCM). The concentrations of interferon-gamma (IFN-gamma), interleukin-2 (IL-2), IL-4, and IL-10 in splenocyte culture supernatants were detected by enzyme-linked immunoabsorbent assay (ELISA), and mRNA levels of the cytokines in splenocytes were assayed by reverse transcription-polymerase chain reaction (RT-PCR).The proliferation of splenocytes and mortality of H22 cells were significantly higher in RF group than in surgical resection group, normal control group, and tumor-bearing group (P0.05). The concentration and mRNA level of IL-2 and IFN-gamma were significantly higher in RF group than in surgical resection group, normal control group, and tumor-bearing group (P0.05); the concentration and mRNA level of IL-4 and IL-10 were significantly lower in RF group than in surgical resection group, normal control group, and tumor-bearing group (P0.05). No significant difference were found between surgical resection group and normal control group (P0.05).RF can efficiently stimulate splenocyte activation and proliferation in H22-bearing mice, and enhance splenocyte cytotoxicity to tumor cells. RF could facilitate the secretion and gene transcription of Th1 type cytokines in H22 cells.