To investigate the expression intensity and prognostic significance of TGF-β1 protein in non-small cell lung cancer (NSCLC), immunohistochemistry was carried out in 194 cases of NSCLC and 24 cases of normal lung tissues by SP methods. The PU (positive unit) value was used to assess the TGF-β1 protein expression in systematically selected fields under the microscope with Leica Q500MC image analysis. We found that the TGF-β1 PU value was nearly two-fold higher in NSCLC than in normal lung tissues (p=0.000), being associated with TNM stages (p=0.000) and lymph node metastases (p=0.000), but not to patient age, gender, smoking history, tumor differentiation, histological subtype and tumor location (P>0.05). Univariate analysis indicated that patients with high TGF-β1 protein expression and lymph node metastases demonstrated a poor prognosis (both p=0.000, ). Multivariate analysis showed that TGF-β1 protein expression (RR = 2.565, p=0.002) and lymph node metastases (RR=1.874, p= 0.030) were also independent prognostic factors. Thus, TGF-β1 protein expression may be correlated to oncogenesis and serve as an independent prognostic biomarker for NSCLC.