Adaptive optics enables multimode 3D super-resolution microscopy via remote focusing

A variety of modern super-resolution micro-scopy methods provide researchers with previouslyinconceivable biological sample imaging opportunities ata molecular resolution. All of these techniques excel atimaging samples that are close to the coverslip, howeverimaging at large depths remains a challenge due to aber-rations caused by the sample, diminishing the resolution ofthe microscope. Originating in astro-imaging, the adaptiveoptics (AO) approach for wavefront shaping using adeformable mirror is gaining momentum in modern mi-croscopy as a convenient approach for wavefront control.AO has the ability not only to correct aberrations but alsoenables engineering of the PSF shape, allowing localiza-tion of the emitter axial position over several microns. Inthis study, we demonstrate remote focusing as another AO benefit for super-resolution microscopy. We show theability to record volumetric data (45×45×10 μm), whilekeeping the sample axially stabilized using a standard widefield setup with an adaptive optics addon. We processed the data with single-molecule localization routines and/or computed spatiotemporal correlations, demonstrating subdiffraction resolution.