Optimized approach for optical sectioning enhancement in multifocal structured illumination microscopy

Multifocal structured illumination microscopy (MSIM) is the parallelised version of image scanning microscopy (ISM) by using many excitation spots, which provides a two-fold resolution enhancement beyond the diffraction limit with a frequency of 1 Hz per 3D picture, but scattered and out-of-focus light in thick samples degrades MSIM optical sectioning performance. Herein, we introduce a new optical sectioning method in MSIM via illumination fluctuation. The proposed method suppresses the out-of-focus light by taking full advantage of the statistic property of MSIM raw data, has no requirement of changing system setup or projecting more illumination patterns. Experimental results demonstrate that the method can reduce the out-of-focus light by 7.25 times in optical sectioning image.