The presence of macrophages resulted in at least a threefold upregulation in the release of vascular endothelial growth factor (VEGF) in vitro when compared with spheroids composed only of tumour cells. A homogeneous distribution of macrophages surrounding the hypoxic centre was observed in the majority of spheroid sections assessed. The angiogenic response measured around the spheroids 3 days after in vivo implantation was significantly greater in the spheroids infiltrated with macrophages; the number of vessels increased (macrophages vs no macrophages, 34 ± 1.9 vs 26 ± 2.5, P < 0.01), and were shorter in length (macrophages vs no macrophages, 116 ± 4.92 vs 136 ± 6.52, P < 0.008) with an increased number of junctions (macrophages vs no macrophages, 14 ± 0.93 vs 11 ± 1.25, P < 0.025), all parameters indicative of new vessel formation. By day 7 no significant differences were seen. Viable human but no murine macrophages were identified in the tumour spheroids at the end of the study, using immunohistochemistry.