We explored the mechanisms of class B CpG-oligodeoxynucleotide-induced antitumour effects against weakly immunogenic tumours. Treatment with CpG-oligodeoxynucleotide 1826 (CpG) induced similar antitumour effects in B16 melanoma-bearing immunocompetent C57BL/6 mice and T-cell-deficient severe combined immunodeficient (SCID) mice, and NXS2 neuroblastoma-bearing T-cell-depleted A/J mice. Both macrophages (Mφ) and natural killer (NK) cells from CpG-treated C57BL/6 mice could mediate cytotoxicity in vitro, suggesting that these cell types might control tumour growth in vivo. However, CpG treatment of SCID/beige mice or T-cell-depleted and NK-cell-depleted A/J mice still induced antitumour effects in vivo, arguing against a major role of NK cells in the antitumour effects of CpG in the absence of T cells. In contrast, CpG treatment of interferon-γ knockout (IFN-γ–/–) C57BL/6 mice resulted in no antitumour effects in vivo and no Mφ-mediated tumoristasis in vitro despite unaltered cytolytic function of NK cells in vitro. Moreover, Mφ inactivation by silica substantially reduced CpG-induced suppression of tumour growth in vivo, revealing an important role of Mφ in CpG-induced antitumour effects. The in vitro tumouritoxicity by CpG-stimulated Mφ (CpG-Mφ) correlated with tumour cell mitochondria dysfunction and involved nitric oxide (NO), tumour necrosis factor-α (TNF-α) and IFN-γ, whereas interleukin-1α (IL-1α), IL-1β, IFN-α, TNF-related apoptosis-inducing ligand and Fas ligand played insignificant roles in CpG-Mφ tumouritoxicity. Taken together, our results indicate that the growth control of weakly immunogenic tumours during CpG-immunotherapy is mediated predominantly by Mφ, rather than T cells or NK cells.