Single-objective selective-volume illumination microscopy enables high-contrast light-field imaging
Sara Madaan, Kevin Keomanee-Dizon, Matt Jones, Chenyang Zhong, Anna Nadtochiy, Peter Luu, Scott E. Fraser, Thai V. Truong
Abstract
The performance of light-field microscopy is improved by selectively illuminating the relevant subvolume of the specimen with a second objective lens. Here we advance this approach to a single-objective geometry, using an oblique one-photon illumination path or two-photon illumination to accomplish selective-volume excitation. The elimination of the second orthogonally oriented objective to selectively excite the volume of interest simplifies specimen mounting; yet, this single-objective approach still reduces the out-of-volume background, resulting in improvements in image contrast, effective resolution, and volume reconstruction quality. We validate our new, to the best of our knowledge, approach through imaging live developing zebrafish, demonstrating the technology's ability to capture imaging data from large volumes synchronously with high contrast while remaining compatible with standard microscope sample mounting.