Although the clinical use of vincristine is limited by its adverse effect, neuropathic pain, the mechanism of this effect is poorly understood. Recently, reports demonstrated that inflammatory and immune responses play an important role in the neuropathic pain that follows peripheral nerve injury. In this study, we examined the role of macrophage-derived interleukin (IL)-6 in vincristine-induced mechanical allodynia. Vincristine sulfate (0.01–0.1 mg/kg, i.p.) was administered to male ICR mice and BALB/c mice once per day for 7 or 14 days. Mechanical allodynia was evaluated by withdrawal responses, using von Frey filaments from day 0 to day 28. In both ICR mice and BALB/c mice, significant dose-dependent increases in the percentage of withdrawal responses were observed from day 3 to day 28 following repeated administration of vincristine (0.1 mg/kg). As determined by immunohistochemistry, the number of macrophages in the region of the sciatic nerve and lumbar dorsal root ganglion was significantly increased on day 7 of vincristine administration. The expression of IL-6 was increased by vincristine administration and was co-localized in the invading macrophage. Moreover, a neutralizing antibody of IL-6, which was injected into areas surrounding the sciatic nerve on day 0, 3, and 6, significantly attenuated vincristine-induced mechanical allodynia from day 7 to day 28. In addition, the incidence of vincristine-induced mechanical allodynia in IL-6 knockout mice was lower than that in wild type mice from day 3 to day 28. These results suggest that the invading peripheral macrophage-derived IL-6 plays a critical role in vincristine-induced mechanical allodynia.