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

Abstract

fluorescence microscopy with sensor-based conjugate adaptive optics using oblique back illumination," J. 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.

Fourier conjugate adaptive optics for deep-tissue large field of view imaging

“…As a result, conjugate AO allows the use of part of the previously optimized wavefront during the scanning procedure and provides a significant FOV advantage [22,23]. The efficiency of this approach was demonstrated in recent experiments [16,[24][25][26][27].…”

Single-shot quantitative aberration and scattering length measurements in mouse brain tissues using an extended-source Shack-Hartmann wavefront sensor

“…Direct wavefront sensing for AO-enhanced optical microscopy has been reported over the last years through several approaches, using either Shack-Hartmann (SH) sensors [13-16, 22, 23], or partitioned aperture wavefront (PAW) sensors [24] as a variant of pyramid wavefront sensors applicable to microscopy. The use of PAW in fluorescence microscopy is severely limited, since it requires only a moderate spatial incoherence of the excitation source, so that its demonstration in fluorescence microscopy was restricted to widefield using a specific illumination geometry (Oblique Back Illumination) [24,25]. Direct wavefront sensing in microscopy is thus currently mostly based on SH sensors.…”