Abstract Background Monocyte recruited into the tumor and maturation to tumor-associated macrophage (TAM). Interleukin-10(IL-10) is a potent immunosuppressive cytokine, which can be secreted from both primary tumor and stromal cells. However, there are controversies regarding its role in the progression of cancer. So it is important to isolate TAM from tumor cells to study the role of IL-10 in the progress of cancer. The aim of our study was to determine whether IL-10 expressed by TAM correlated with clinicopathological factors in NSCLC. Methods TAM in NSCLC was isolated by short-term culture in serum free medium with the modification to literature reports. The mRNA expression levels of IL-10, cathepsin B, cathepsin S, which were closely related with TAM according to the literatures, were evaluated by Quantitative real-time RT-PCR in 63 NSCLC. The relationships between their expression levels and clinicopathological features were investigated. Results We successfully achieved up to 95% purity of TAM, derived from 63 primary lung cancer tissues. TAM expressed high levels of IL-10, cathepsin B in NSCLC. High levels of IL-10 in TAM significantly correlated with stage, tumor size, lymph node metastasis, lymphovascular invasion or histologic poor differentiation. Conclusions Our results revealed that TAM with high levels of IL-10 expression may play an important role in the progression of non-small cell lung cancer. The data also suggested that TAMs may involve in tumor immunosuppression through overexpressed IL-10. Additionally, the phenotype of isolated TAM can be potentially used to predict clinicopathological features as well.