High-Resolution Line-Scanning Two-Photon Microscope

A two-photon fluorescence microscope employing line-shaped illumination is presented. This type of excitation is commonly expected to bring about the degradation of axial resolution because of the weaker focusing of the illuminating beam in just one direction. On the basis of a detailed theoretical investigation of the beam shaping performed by cylindrical lenses when inserted in conventional point-scanning systems, we design and implement a microscope set-up making use of readily available optical components. The experimental results show that the proper choice and arrangement of the cylindrical lenses that we have devised is able to preserve the optical-sectioning capability at the video-rate acquisition speed.