Amplitude-modulation-type multi-ring mask for two-photon excitation scanning microscopy

Abstract

A smaller point spread function (PSF) is desirable for high-resolution image acquisition. In two-photon excitation microscopy, the focus size of the excitation light is directly related to the two-photon fluorescence PSF. To achieve a small PSF, we designed amplitude-modulation-type multi-ring pupil masks using two design methods. When the intensity distribution of the excitation light has a top-hat form, the volume of the PSF obtained with the first designed multi-ring mask is 1.46 times smaller than that obtained without the mask. Furthermore, the second multi-ring mask is less influenced by spherical aberration occurring during observation inside the sample. Our method, which is robust against spherical aberration, reduces the size of the PSF in the observation of fluorescent beads and biological samples.