Influences of aberration on spatial resolution of STED microscope in probing a specimenwith discontinuous refraction indices.

Probing depth and system aberrations have direct impacts on the spatial resolution of stimulated emission depletion (STED) microscopes. Based on the vectorial diffraction theory, the influence of coma and astigmatism on the focal patterns of STED microscopes in probing stratified mediums with discontinuous refractive indices (e.g., glass cover slip, solution, and biological samples, etc.) have been illustrated in detail. The spatial resolution of the STED system has been discussed by analyzing the full width at half-maximum size of the fluorescence spots. It is found that, while probing in stratified media with discontinuous refractive indices, the spatial resolution of a STED microscope can be very sensitive to the existence of aberrations, e.g., coma and astigmatism, at different probing depths, as a result of mismatched axial positions of the excitation and depletion patterns. The spatial resolution of STED can be degraded up to 1.87- and 1.95-fold compared to that without aberrations. Therefore, a careful evaluation of the influence of aberration and discontinuous refractive indices should be taken into account when applying a STED microscope to realize super-resolution images.