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Background and objective Due in part to remarkable advances over the past decade in our understanding of lung cancer, there is a pressing need for a revised classification. The aim of this study was to explore and verify the clinical significance of a new molecular classification of non-small cell lung cancer (NSCLC) on the basis of anatomic site of bronchial tree. Methods Double-immunohistochemistry were performed in 105 tumor samples paring with normal lung tissue from non-small cell lung cancer patients by using monoclonal antibody of P63/NapsinA and TTF-1/CK7. By comprehensively analyzing the express profiles of tumors and normal lung tissues, histological characteristics we proposed a brand new pathological classification based on histogenesis for NSCLC and divided them as: bronchiole epithelial cell carcinoma, bronchiole-alveolar cell carcinoma, alveolar cell carcinoma and secretory adenocarcinoma. And to analyze the relationship between this classification and epidermal growth factor receptor (EGFR) mutation and ognosis. Results Further investigation revealed that our new classification showed strong relevancy with EGFR mutations and effective indicators for prognosis. Conclusion The classification system for NSCLC proposed by our research group is convenient for pathological diagnosis and valuable in clinical application.