Purpose: To assess noninvasive optical coherence tomography (OCT) and optical Doppler tomography (ODT) for early detection and evaluation of chemotherapy-induced oral mucositis. Experimental Design: Cheek pouches of 10 Syrian golden hamsters were imaged using OCT/ODT during development of chemotherapy-induced mucositis. I.p. injections of 5-fluorouracil and mechanical irritation induced oral lesions. At 2, 4, 7, and 11 days, one hamster was sacrificed and processed for histopathology. OCT images were visually examined; ODT results were semiquantified. Imaging data were compared with histologic findings. Results: During the development of mucositis, OCT/ODT identified the following events: (a) change in epithelial thickness (beginning on day 2), (b) loss of surface keratinized layer continuity (beginning on day 4), (c) loss of epithelial (day 4 onwards) and submucosal integrity (day 7 onwards), (d) changes in axial blood flow velocity (increased on days 2 and 4; decreased on day 7), and (e) changes in blood vessel size (diameter doubled on day 2; quadrupled on day 4; unchanged on day 7). The semiquantitative imaging-based scoring system identified the severity of mucositis as defined by histopathology. The combination of imaging criteria used allowed for the detection of early, intermediate, and late mucositic changes. Imaging data gave higher scores compared with clinical scores early on, suggesting that the imaging-based diagnostic scoring was more sensitive to early mucositic change than the clinical scoring system. Once mucositis was established, imaging and clinical scores converged. Conclusion: OCT/ODT identified chemotherapy-induced oral changes before their clinical manifestation, and the proposed scoring system for oral mucositis was validated for the semiquantification of mucositic change. © 2007 American Association for Cancer Research.