Wavefront shaping of a Bessel light field enhances light sheet microscopy with scattered light

Abstract

Light sheet microscopy has seen a resurgence as it facilitates rapid, high contrast, volumetric imaging with minimal sample exposure. Initially developed for imaging scattered light, this application of light sheet microscopy has largely been overlooked but provides an endogenous contrast mechanism which can complement fluorescence imaging and requires very little or no modification to an existing light sheet fluorescence microscope. Fluorescence imaging and scattered light imaging differ in terms of image formation. In the former the detected light is incoherent and weak whereas in the latter the coherence properties of the illumination source, typically a laser, dictate the coherence of detected light, but both are dependent on the quality of the illuminating light sheet. Image formation in both schemes can be understood as the convolution of the light sheet with the specimen distribution. In this paper we explore wavefront shaping for the enhancement of light sheet microscopy with scattered light. We show experimental verification of this result, demonstrating the use of the propagation invariant Bessel beam to extend the field of view of a high resolution scattered light, light sheet microscope and its application to imaging of biological super-cellular structures with sub-cellular resolution. Additionally, complementary scattering and fluorescence imaging is used to characterize the enhancement, and to develop a deeper understanding of the differences of image formation between contrast mechanisms in light sheet microscopy.