Objective: Well-differentiated papillary adenocarcinoma of the lung (G1 cancer cells) is difficult to distinguish from benign bronchial columnar epithelial cells with reactive atypia (benign cells) in many cases because nuclear atypia is mild. We focused on the 3-dimensional presence of nuclei in cell smears. Several images focused on the nucleus were acquired, and the nuclear luminance was measured and analyzed. Study Design: One hundred G1 cancer cells and benign cells (nuclei), respectively, were selected from those on a bronchial brushing preparation for cytology. Images of 41 layers were acquired at 0.25-µm intervals in each cell, and the nuclear luminance was measured (a total of 8,400 images). Results: There were more focus positions in the G1 cancer cell nuclei, showing a 3-dimensional nucleus, compared to benign cells, and the 3-dimensional variation in the coefficient of variation (CV) of nuclear luminance at the focus position was smaller in G1 cancer than in benign cells, showing a significant difference. Conclusion: The G1 cancer cells’ nuclear structure was more 3-dimensional, and the chromatin distribution was homogeneous. The three-dimensional variation in the nuclear luminance CV could be numerically presented, which might be an objective index for cancer diagnosis.