TIME-RESOLVED AND WAVELENGTH-RESOLVED SPECTROSCOPY IN 2-PHOTON-EXCITED FLUORESCENCE MICROSCOPY

Two-photon excited fluorescence spectroscopy has been performed at a m icroscopic scale in combination with normal, white-light microscopy. T his gave simultaneously a spectral resolution of 20 nm and a temporal resolution of 20 ps, from a volume element less than 5 mu m in all thr ee dimensions. The sample was excited with the light from a continuous ly mode-locked Ti:sapphire laser that was focused on the sample in a f luorescence microscope. A polychromator and a streak-camera were used for detection. The method has been used on tissue, plant and paper sam ples. It has also been demonstrated how substances naturally occurring in the samples can be identified from their spectroscopic properties and the spatial distribution of these substances can be observed.