Improving the space-bandwidth product of structured illumination microscopy using a transillumination configuration

Abstract

Applying structured instead of plane illumination in widefield optical fluorescence microscopy can improve the spatial resolution beyond what is known as the Abbe limit. In general it is not only the resolution of an imaging system that is of interest but also its field of view (FOV). These two parameters are expressed in the space-bandwidth product (SBP). Here we introduce a modified structured illumination microscopy (SIM) approach that offers a larger SBP than any other available implementation. This is achieved through a transillumination geometry instead of the typical epifluorescent configuration. Compared to conventional SIM, the illumination path is decoupled from the objective lens by using a multi-mirror setup to generate the sinusoidal interference pattern for structured illumination in transmission mode. The spatial frequency of the illumination pattern can be controlled by changing the angle of the mirrors, achieving comparably fine patterns over a large FOV. In this work simulation results demonstrate the potential resolution improvement to be expected by the suggested implementation. Preliminary experimental results demonstrate phase-shifting ability and the stability of fringe frequencies over a large FOV of (≈16 mm²) at different numerical apertures, fulfilling the prerequisites for SIM acquisition.