Mechanism of STED microscopy and analysis of the factors affecting resolution

摘要

Stimulated emission depletion (STED) microscopy exploits nonlinear saturable optical transition of fluorescent molecules, allowed to overcome Abbe's diffraction-limit and provides diffraction-unlimited resolution in far-field optical microscopy. We elaborate the mechanism of STED and the conditions of depletion. The formula of STED microcopy resolution is deduced through effective point spread function (E-PSF). The STED system resolution is mainly dominated by the quality of the fluorescence depletion patterns in the focal plane. The depletion pattern is mainly affected by STED beam intensity, polarization, phase plate, primary aberrations, STED pulse shape, pulse duration and delay time. In this paper, we found related models and simulate the relationship between the depletion patterns and the parameters, and put forward effective approach to enhance the system resolution.