Simulating structured-illumination microscopy in the presence of spherical aberrations

The effect of depth-induced spherical aberrations (SA) on structured illumination microscopy (SIM)1 is investigated. SIM is a technique used in three-dimensional (3D) fluorescence microscopy to improve resolution in optical sections acquired from 3D specimens. A 3D depth-variant imaging model was developed to predict the intermediate SIM or grid images that are used by the SIM approach to compute improved optical sections. The model incorporates SA due to imaging depth within a sample when there is a refractive index (RI) mismatch between the average RI of the specimen and the RI of the immersion medium of the lens. The model was implemented using a stratum-based model approximation and multiple depth-variant point-spread functions (PSFs) 2. SIM optical sections were computed using the subtraction algorithm 1,3 and simulated grid images that include SA predicted by our model. Simulations were performed for different imaging conditions by varying the grid frequency, the amount of SA and the level of noise added to the grid images. Simulated results demonstrate that SIM images are less accurate in the presence of SA, and confirm that the SIM approach is very sensitive to system noise resulting in a reduced SNR in the optically sectioned images. © 2011 SPIE