The aim of this study was to examine the ability of endobronchial ultrasonography (EBUS) to image the bronchial wall structure in order to assess the depth of bronchial tumor invasion. Sixty-one patients who underwent lobectomy, pneumonectomy or forceps biopsy were included in this study. In 21 patients with bronchoscopically visible bronchial malignant tumors, EBUS was performed during bronchoscopy. In the remaining 40 patients, ultrasonography was performed on the resected specimens. The EBUS findings obtained using thin ultrasonic probes (20 MHz radial scanner) were compared with the macroscopic and histologic findings of the corresponding areas in the resected specimens. When the bronchial walls were imaged while immersed in normal saline, six ultrasonically distinct layers were detected in the cartilaginous and membranous portions. A similar wall structure was imaged when EBUS was performed during bronchoscopy using a latex balloon sheath. The image of the lamina propria and submucosa was occasionally compressed and mixed with a balloon echo due to the latex balloon sheath, whereas the cartilage layer was always distinctly imaged. A good correlation was observed between the EBUS-determined cartilage thickness and the actual histologic measurement, as measured with vernier calipers. Malignant tissues were imaged as hypoechoic areas, and tumor invasion of the cartilage layer was clearly detected. In conclusion, using high-resolution (20 MHz) ultrasonic probes, the bronchial wall structure could be imaged as six distinct layers. The cartilage layer was easily identified and could be used as a reference to evaluate the rest of the bronchial wall structure.