Boosting the performance of light-sheet microscopy via synchronous detection from the illumination and detection light path

Abstract

The image quality of light-sheet microscopy degrades due to the system misalignment or opacity of the sample. In this work, we proposed to synchronously detect the fluorescence from both the illumination and detection light path of axially swept light-sheet microscopy (SD-LSM) to realize the full exploitation of the excited fluorescence. We adopted spatially variable multi-view deconvolution to fuse images from the detection and illumination objective of SD-LSM to improve the resolution degradation caused by the nonlinearity of scanning devices. We proposed the fusion of images from the detection and illumination objective of SD-LSM based on background estimation to improve the signal-to-background ratio (SBR). We separately demonstrated that the spatial resolution and the SBR can be largely boosted by SD-LSM for various biological samples, after the fusion of images from the illumination and detection path. Compared with the images only from the detection path, images from SD-LSM showed the axial resolution recovery by up to 14.6 times when axial scanning devices work at high speed with large nonlinearity, and SBR enhancement by up to 8.2 dB when imaging a highly scattered sample. SD-LSM could boost the image quality without any additional time consumption for image acquisition or photon budget for the sample at a cost of a simple addition of a camera in the illumination path, compared with conventional axially swept light-sheet microscopy.