Adaptive speckle illumination microscopy

Wide field microscopy is well established in biological and medical applications. However, its reduced depth sectioning capability leads to background signals originating outside the depth of interest that degrade the contrast and limit the usability. Confocal microscopy is advantageous due to its optical sectioning, but it requires scanning in three dimensions to obtain 3D information, which is time consuming and may result in photobleaching. Hybrid illumination microscopy combines the advantage of both worlds: it is camera based and uses speckle illuminations to introduce optical sectioning capabilities. A fast digital mirror device (DMD) is employed for speckle modulation. We present adaptive lens-based axial scanning for volumetric measurements in hybrid illumination microscopy. Successful volumetric measurements of fluorescent thyroid glands in zebrafish embryos underline the potential of this approach. Perspectives of various motion-free illumination approaches as well as novel adaptive lenses with tunable asphericity are discussed.