Purpose: To determine optimal imaging time for diagnostic I-123 whole body scan in the follow-up of patients with differentiated thyroid cancer (DTC), we compared the image quality of 6- and 24-hour images of the same subjects. Materials and Methods: Four hundred ninety-eight patients (M:F=55:443, Age 47.6¡¾12.9 years) with DTC who had undergone total thyroidectomy and I-131 ablation therapy underwent diagnostic whole body scanning 6 hour and 24 hour after oral ingestion of 185 MBq (5 mCi) of I-123. Serum thyroglobulin measurement and ultrasonography of the neck were performed at the time of imaging. In 40 patients underwent additional I-131 therapy, post-therapy I-131 images were obtained and compared with diagnostic I-123 images. Results: In 440 patients (88.4%), 6- and 24-hour diagnostic I-123 images were concordant, and 58 patients (11.6%) showed discordant findings. Among 58 discordant patients, 31 patients showed abnormal tracer uptake on only 6-hour image, which turned out false-positive findings in all cases. In 12 patients with positive findings on only 24-hour image, remnant thyroid tissue (4 patients) and cervical lymph node metastasis (3 patients) were presented. Among 40 patients underwent additional I-131 therapy, 6-hour and 24-hour images were discordant in 13 patients. All 5 patients with abnormal uptake on only 6-hour image revealed false-positive results, whereas most of 24-hour images were concordant with post-therapy I-131 images. Conclusion: I-123 imaging at 24-hour could reduce false-positive findings and improve diagnostic accuracy, compared with 6-hour image in the follow-up of patient with DTC. (Nucl Med Mol Imaging 2009;43(2):129-136)