Abstract

Confocal laser scanning microscopy provides a significant improvement in axial resolution over conventional epi-fluorescence microscopy by eliminating out of focus fluorescence using a spatial filter in the form of a confocal aperture Combining two-photon induced fluorescence with laser scanning microscopy, Denk et al. [1] achieved high axial/depth discrimination even without a confocal aperture in front of the photodetector, due to the quadratic dependence of the two-photon absorption on the laser intensity Strong fluorescence is only induced in the vicinity of the focal point The background scattering light and autofluorescence of the sample in the two-photon excited system is also lower. The use of infrared wavelength leads to deeper penetration depth in most materials, providing the opportunity to image thicker samples Two-photon scanning microscopy have been applied extensively in the field of biology and biomedicine, taking advantage of the recent rapid development and commercial availability of femtosecond Ti:sapphire lasers However, recent studies on human skin and biological tissues have suggested that the highest penetrating infrared wavelength in most human and biological tissues exists around 1200-1250 nm, which is not available with Ti:sapphire lasers

© 2000 IEEE