Effects of aberrations on isoSTED microscopy

Abstract

Aberrations are inevitable in optical microscopes and affect the image quality. In STED microscopy, the optical resolution is especially dominated by the performance of depletion pattern. To quench fluorescence only in the periphery of the excitation focus, the depletion focus ideally has a zero-intensity point that is surrounded by a high-intensity crest. Otherwise, distortion of the depletion beam causes unwanted fluorescence inhibition. The influence of aberrations on isoSTED microscopy differs from that on STED microscopy because isoSTED microscopy generates a more complicated shape of the depletion focal spot with two opposing objectives which have different focusing properties. We present analysis that elucidates the effects on the shape, the central minimum intensity, and the peak intensity of the isoSTED pattern regarding primary aberrations such as astigmatism, coma, and spherical aberration when those aberrations are added in the shared beam path and each beam path in the 4Pi-cavity. The simulation results demonstrate significant resolution degradation (>160%; defined as the focal volume) for the aberrations with root mean square wavefront errors of 0.5 rad.