Ki-67 and proliferating cell nuclear antigen (PCNA) are markers of proliferation used to assess the growth fraction of the cell population. The present study aimed to explore the prognostic value of these proliferative markers in patients with resected esophageal squamous cell cancer (ESCC) in a large cohort. A total of 807 patients with ESCC who underwent radical resection were retrospectively reviewed. Ki-67 and PCNA index were examined as the percentage of positively nuclear-stained cells among total number of cancer cells in three high-power fields by a pathologist who was blinded to the patients' history and outcome. Overall survival (OS) and disease-free survival (DFS) were estimated. The Cox regression model was used to evaluate the independent factor. The cut-off value as 60 and 80% for Ki-67 and PCNA were verified, respectively. Higher Ki-67 expression was associated with low differentiation and more lymph node metastasis. Higher PCNA expression was associated with elevated T stage. However, either expression of Ki-67 or PCNA was not correlated with OS and DFS. While in combination of Ki-67 and PCNA analysis, higher expression of these two proliferative markers predicted worse prognosis (median OS, 47 months versus 54 months, P = 0.04). Whatever the combined proliferative marker, differentiation, lymph node metastasis stage and vascular invasion act as factors in univariate survival analysis, but combined Ki-67 and PCNA is not an independent prognostic variable in multivariate analysis (P = 0.10). Our results suggest that proliferative markers of Ki-67 and PCNA may correlate with tumor stage but cannot act as independent predictor of prognosis in ESCC patients.