Nodal staging is a crucial factor in choosing the treatment option for non-small cell lung cancer (NSCLC). However, so far as we know, a computer-based histomorphometric analysis of lymph nodes in NSCLC has never been developed. We studied 299 surgically resected lymph nodes from 108 patients with NSCLC. Microscopic digital images were analyzed with Scion Image software. Seventy lymph nodes had at least one metastatic focus. The metastasis occupancy area per node ranged from 0.01 to 209.58 mm(2) (mean, 29.58 +/- 5.87 mm(2)). The metastasis occupancy ratio ranged from 0.01% to 100% (mean, 48.70% +/- 42.10%). The short-axis diameter of malignant lymph nodes was significantly longer than that of benign lymph nodes (P = 0.0002). The average metastasis occupancy area in the regional lymph nodes of NSCLC is quite small. Various inflammatory conditions can result in a false-positive diagnosis when these techniques are used. Studies that combine analysis of primary tumor size and serum carcinoembryonic antigen (CEA) levels with imaging methods should be considered. Finally, the use of mediastinoscopy or video-assisted thoracoscopic surgery is encouraged in determining the exact nodal status in NSCLC.