PROCEEDINGS ARTICLE | October 27, 2006
Proc. SPIE. 6047, Fourth International Conference on Photonics and Imaging in Biology and Medicine
KEYWORDS: Confocal microscopy, Second-harmonic generation, Cornea, Tissues, Skin, Luminescence, Iris, Sclera, Collagen, Harmonic generation
We report the implementation of compound imaging with backward second harnionic generation (SHG) and two-photon
excitation fluorescence (TPEF) in a standard confocal microscope. Various biological tissues, including skin, muscle and
ocular tissues (e.g. cornea. iris, choroid, and sclera) have been investigated with different sample preparation methods
(fresh, fixation). The results show that collagen-rich ocular tissues, like cornea, iris, choroids, and sclera can produce
strong SHG signals. In sclera, there are not only plenty of collagen, but also various endogenous fluorophores. The
organization of collagen fibers in sclera is clearly distinguishable from its SHG images viewed transversely and
horizontally respectively. We also find that fixation of the sample with formaldehyde has reduced the intensity of SHG
intensity by almost 5O%. As the SHG and TPEF can provide complementary information about collagen and
fluorophores rich biological tissues, compound imaging of SHG and TPEF presented in this study is believed to have
potential applications in biomedicine and clinical diagnosis.
