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Ray-tracing study on the post-scanner variable beam expansion optics in a two-photon microscopy system

Due to the low signal levels typical of two-photon microscopy (TPM) in biological samples, optical design optimization is critical. One of the most important factors is overfilling of the back aperture of the objective lens. A variable beam expander is commonly placed before the scanning mirrors to achieve this goal, however, this may cause degradation of image quality due to increased dispersion. Additionally, scanning mirror size restricts the degree of expansion, which often prevents the overfilling of objective lens back aperture. We investigated the implementation of variable beam expansion optics after the scanning mirrors. Ray-tracing analyses confirmed that the post-scanner beam expansion has two key advantages over the conventional pre-scanner beam expansion approach: decreasing the number of optical elements reduces pulse dispersion and reducing the size of the scanning mirror enables faster scanning. Resolution and aberration of a TPM with post-scanner beam expansion optics were analysed.