Light sheet microscopy with high spatial resolution based on polarized structured illumination beam modulated by the phase mask

Abstract

In this article, we present a method to improve axial resolution and expand the field of view of conventional light sheet fluorescence microscopy (LSFM) by phase modulation of the illumination beam. We added a bisected annular binary phase mask on the illumination path to modulate the radially polarized beam. The central spot of the modulated point spread function is narrower than that of Gaussian focus, although the sidelobe energy is increased. Then, a diffraction grating is used to produce multiple copies of this illumination pattern, and these pattern copies form the light sheet. By matching the numerical apertures of illumination lens and detection lens, the sidelobes on the detection axis can be suppressed in the captured data, although they do still contribute to photobleaching and phototoxicity in the sample. The simulation results show that the proposed method effectively reduces the thickness of the light sheet by 1.3 times, and increase the length of the light sheet by two times compared with conventional LSFM.

Introduction to the special section on light sheet fluorescence illumination microscopy

“…In particular, electrically tunable lens were proven to be an optimal solution to improve axial resolution maintaining the minimal light sheet thickness over a wide field of view (Hedde & Gratton, ). Furthermore, methods able to improve axial resolution and simultaneously expand the field of view through phase modulation, obtained by modulating the radially polarized beam with annular binary phase mask (Nhu, Wang, Liu, Kuang, & Liu, ).…”