Two-photon laser scanning fluorescence microscopy-from a fluorophore and specimen perspective

Abstract

Two-photon laser scanning microscopy is discussed from a fluorophore and specimen point of view. Issues to be considered in the selection of the fluorophore and in specimen preparation for multiphoton microscopy are described. The transverse resolution is estimated, based on a photobleaching method. We also show that the performance of a multiphoton laser scanning microscope is significantly enhanced by the use of a confocal aperture.