High-NA open-top selective-plane illumination microscopy for biological imaging

Abstract: Selective-plane illumination microscopy (SPIM) provides unparalleled advantages for volumetric imaging of living organisms over extended times. However, the spatial configuration of a SPIM system often limits its compatibility with many widely used biological sample holders such as multi-well chambers and plates. To solve this problem, we developed a high numerical aperture (NA) open-top configuration that places both the excitation and detection objectives on the opposite of the sample coverglass. We carried out a theoretical calculation to analyze the structure of the system-induced aberrations. We then experimentally compensated the system aberrations using adaptive optics combined with static optical components, demonstrating near-diffraction-limited performance in imaging fluorescently labeled cells. © 2017 Optical Society of America OCIS codes: (080.080) Geometric Optics; (110.0110) Imaging systems; (110.0180) Microscopy.