Tiling light sheet selective plane illumination microscopy using discontinuous light sheets.

Abstract

Tiling light sheet selective plane illumination microscopy (TLS-SPIM) improves the 3D imaging ability of SPIM by using real-time optimized tiling light sheets. However, the imaging speed decreases and the raw image size increases due to the tiling process and additional camera exposures. The decreased imaging speed and the increased raw data could cause significant problems when TLS-SPIM is used to image large specimens at high spatial resolutions. Here, we present a novel method to solve the problem. Discontinuous light sheets created by scanning coaxial beam arrays synchronized with the detection camera rolling shutter are used in TLS-SPIM for 3D imaging. It improves the imaging efficiency of TLS-SPIM by reducing the number of tiles required per image plane without influencing the spatial resolution. We investigate the method via numerical simulations and experiments. We demonstrate the imaging ability of the TLS-SPIM using discontinuous light sheets and show the improved imaging efficiency by imaging optically cleared mouse brain.