High-speed combined reflectance confocal and moxifloxacin based two-photon microscopy

Reflectance confocal microscopy (RCM) is a non-invasive high-resolution optical imaging technique used in clinical settings as a diagnostic method. However, RCM has limited diagnostic ability by providing non-specific morphological information only based on reflection contrast. Various multimodal imaging techniques have been developed to compensate the limitations of RCM, but multimodal techniques are often slow in imaging speed compared to RCM alone. In this report, we combined RCM with moxifloxacin based two-photon microscopy (TPM) for high-speed multimodal imaging. Moxifloxacin based TPM used clinically compatible moxifloxacin for cell labeling and could do non-invasive cellular imaging at 30 frames/s together with RCM. Performance of the combined microscopy was characterized in the imaging of mouse skin and cornea, in vivo. Detail tissue microstructures including cells, extra-cellular matrix (ECM), and vasculature were visualized. The combined microscopy was applied to human skin cancer specimens, and both cells and ECM in the skin cancer and normal skin regions were visualized at high imaging speeds. The combined microscopy can be useful in the clinical applications of RCM by providing multiple contrasts.