The clinical significance of tumor-associated macrophages (TAMs) and CD3-zeta chain expression of tumor-infiltrating lymphocytes (TILs), and their correlation in human esophageal squamous cell carcinoma (SCC) are not very clear. Serial histological slides from 137 esophageal SCC patients who had undergone tumor resection were immunohistochemically studied with anti-CD68, anti-CD3-zeta and anti-CD3-epsilon antibodies. TAMs infiltration (expressed as macrophage index, M(phi)I) and CD3-zeta expression (judged by Z/E = CD3-zeta+ cells/CD3-epsilon+ cells ratio) in different tissue compartments were observed. We found that the total tumor tissue region had significantly higher macrophage density and lower CD3-zeta expression (mean +/- SD: M(phi)I(normal): 225.3 +/- 85.9; Z/E(total): 0.52 +/- 0.25; n = 137) relative to adjacent histologically normal esophageal squamous epithelium (M(phi)I(normal): 60.5 +/- 31.7, P < 0.001; Z/E(normal): 0.79 +/- 0.35, P = 0.001; n = 70). Significantly higher M(phi)I(stroma) (P = 0.006) and lower Z/E(total) (P = 0.016) were detected in patients with lymph node metastasis than in those without. Patients with high M(phi)I(total) and M(phi)I(cancer) but low Z/E(total) had poorer surgical outcomes. Univariate analysis of M(phi)I(total) and multivariate analysis of M(phi)I(total) with specific clinico-pathological parameters demonstrated M(phi)I(total) to be an independent prognostic factor for survival in esophageal SCC patients (Cox proportional hazard model, P = 0.029 and P = 0.031, respectively).