Adaptive optics improves multiphoton super-resolution imaging

Adaptive optics and two-photon instant structured illumination microscopy are combined to provide improved super-resolution imaging within optically aberrating biological samples. We improve multiphoton structured illumination microscopy using a nonlinear guide star to determine optical aberrations and a deformable mirror to correct them. We demonstrate our method on bead phantoms, cells in collagen gels, nematode larvae and embryos, Drosophila brain, and zebrafish embryos. Peak intensity is increased (up to 40-fold) and resolution recovered (up to 176 ± 10 nm laterally, 729 ± 39 nm axially) at depths ∼250 μm from the coverslip surface.