Correcting anisotropic intensity in light sheet images using dehazing and image morphology

Abstract

Light-sheet fluorescence microscopy (LSFM) provides access to multi-dimensional and multi-scale in vivo imaging of animal models with highly coherent volumetric reconstruction of the tissue morphology, via a focused laser light sheet. The orthogonal illumination and detection LSFM pathways account for minimal photobleaching and deep tissue optical sectioning through different perspective views. Although rotation of the sample and deep tissue scanning constitutes major advantages of LSFM, images may suffer from intrinsic problems within the modality, such as light mismatch of refractive indices between the sample and mounting media and varying quantum efficiency across different depths. To overcome these challenges, we hereby introduce an illumination correction technique integrated with depth detail amelioration to achieve symmetric contrast in large field-of-view images acquired using a low power objective lens. Due to an increase in angular dispersion of emitted light flux with the depth, we combined the dehazing algorithm with morphological operations to enhance poorly separated overlapping structures with subdued intensity. The proposed method was tested on different LSFM modalities to illustrate its applicability on correcting anisotropic illumination affecting the volumetric reconstruction of the fluorescently tagged region of interest.