In vivo detection of cryosurgery using multiphoton and harmonic generation microscopy.

The tissue injury and the organization of collagen during cryosurgery are poorly characterized because of the lack of appropriate methodologies. In this study, we use multimodal multiphoton microscopy to assess the change of extracellular matrix after cryotreatment of skin. The cellular matrix transformations and the intercellular interactions during the wound healing process after cryolesion for mice were measured in vivo and in real-time through the dorsal skinfold chamber (DSC). Intrinsic second-harmonic generation (SHG) signals from fibrillar collagen and two-photon excited (TPE) autofluorescence from cell were collected to investigate the cryosurgical response in vivo. The TPE and SHG signals are significantly different between normal and cryotreated mice, and correlates with the wound healing process. The results suggest that this approach may be applied in real-time to noninvasively monitor the cryosurgery process and could potentially be applied to clinical evaluation.