Multiphoton spectral microscopy for imaging and quantification of tissue glycation

Tissue glycation from diabetes and aging can result in complications such as renal failure, blindness, nerve damage and vascular diseases. In this work, we applied multiphoton microscopy for imaging and characterizing the extent of tissue glycation. The characteristic features of multiphoton autofluorescence (MPAF) and second harmonic generation (SHG) images as well as MPAF spectra of glycated bovine skin, cornea and aorta were acquired. The analysis of MPAF intensity change accompanying the glycation process shows that collagen is more responsive to the formation of autofluorescent advanced glycation endproducts (AGE) than elastic fibers. Changes in spectral features were also used to estimate the rate of glycation in tissues with intrinsic AF. Our study shows that multiphton imaging may be used for the in vitro investigation of the effects of tissue glycation and that this approach may be used for monitoring AGE formation in the clinical setting.