Squamous cell carcinoma of the esophagus (ESCC) is one of the most aggressive carcinomas, and it occasionally recurs even in the early stages. Identifying biomarkers that enable stratification of patient survival, especially in the early stages, is important to establish better treatment protocols. By performing microarray analyses, we recently discovered that expression of cytokeratin (CK) 7 and CK14 is related to the clinical outcome of ESCC patients. The aim of the present study was to evaluate the significance of CK7 and CK14 expression in a series of ESCC cases. Tissue sections from 126 surgically resected ESCCs were immunostained for CK7 and CK14, and their expression was evaluated in relation to the clinicopathological findings and outcome. Expression of CK7 and CK14 was detected in 28 (22%) and 106 (84%), respectively, of the 126 ESCCs. CK7-positive was associated with poor differentiation (p=0.049), and CK14-negative was associated with poor differentiation (p<0.001), lymphatic invasion (p=0.027) and advanced stages (p=0.043) of ESCCs. CK7-positive and CK14-negative associated a poor outcome (p=0.014 and 0.013, respectively). Among the stage I/IIA/IIB patients, CK7 expression was a significant predictor of worse survival rate (p<0.001). The 5-year overall survival rate of stage I/IIA/IIB patients with CK7-positive and CK7-negative tumors was 50.0 and 90.3%, respectively. Univariate and multivariate analyses of the stage I/IIA/IIB ESCC patients by the Cox proportional hazards model showed that lymphatic invasion and CK7 expression were significant prognostic factors. Thus, CK7 expression was a useful biomarker for predicting the outcome of stage I/IIA/IIB ESCC.