Bridging the resolution-sectioning gap in STED nanoscopy with coherent-hybrid depletion

Authors:

Michael Belsley, Joana Soares-de-Oliveira, António J. Pereira

Keywords:

axial-lateral resolution; 3D STED; Nijboer-Zernike expansion; PSF; bivortex; phase-based, coherent-hybrid STED

Abstract:

Microscopes generally achieve better 2D imaging compared to 3D, and super-resolution microscopes often aggravate such axial-lateral resolution mismatch. A prime example is vortex phase-based stimulated emission depletion (STED) fluorescence microscopy, which only narrows the point-spread function laterally, thereby increasing the point-spread function (PSF) anisotropy. In this study, we developed a semi-analytical theory based on the Nijboer-Zernike expansion, developed a set of metrics and performed experiments to establish the merits of the alternative, bivortex phase-based, coherent-hybrid STED. We find that this mode reduces the axial-lateral resolution mismatch by providing access to PSF geometries that are strictly forbidden to the two conventional single-beam modes, emulating noisier multi-beam approaches. Both theory and experiment indicate that bivortex STED not only addresses the axial-lateral resolution gap but also delivers a higher signal-to-background ratio than the two more common STED modes that it bridges.

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