Widefield fluorescence microscopy with sensor-based conjugate adaptive optics using oblique back illumination.

Abstract

We describe a wavefront sensor strategy for the implementation of adaptive optics (AO) in microscope applications involving thick, scattering media. The strategy is based on the exploitation of multiple scattering to provide oblique back illumination of the wavefront-sensor focal plane, enabling a simple and direct measurement of the flux-density tilt angles caused by aberrations at this plane. Advantages of the sensor are that it provides a large measurement field of view (FOV) while requiring no guide star, making it particularly adapted to a type of AO called conjugate AO, which provides a large correction FOV in cases when sample-induced aberrations arise from a single dominant plane (e.g., the sample surface). We apply conjugate AO here to widefield (i.e., nonscanning) fluorescence microscopy for the first time and demonstrate dynamic wavefront correction in a closed-loop implementation.