Multimodal nonlinear microscopy of head and neck carcinoma — toward surgery assisting frozen section analysis

BACKGROUND: Treatment of early cancer stages is deeply connected to a good prognosis, a moderate reduction of the quality of life, and comparably low treatment costs. METHODS: Head and neck squamous cell carcinomas were investigated using the multimodal combination of coherent anti-Stokes Raman scattering (CARS), two-photon excited fluorescence (TPEF), and second-harmonic generation (SHG) microscopy. RESULTS: An increased median TPEF to CARS contrast was found comparing cancerous and healthy squamous epithelium with a p value of 1.8·10(-10) . A following comprehensive image analysis was able to predict the diagnosis of imaged tissue sections with an overall accuracy of 90% for a 4-class model. CONCLUSION: Nonlinear multimodal imaging is verified objectively as a valuable diagnostic tool that complements conventional staining protocols and can serve as filter in future clinical routine reducing the pathologist's workload.