179 / 2024-06-14 22:14:27

Fast multiplane multifocal structured illumination microscopy

Super-resolution, SIM, MSIM, fast 3D imaging

Combining the digital pinhole and pixel reassignment method with spot-scanning super resolution microscopy (SRM), multifocal structured illumination microscopy (MSIM) can remove most defocused noise to improve the axial resolution of conventional structured illumination microscopy (SIM), with achieving a two-fold lateral resolution improvement compared to wide-field imaging. However, mechanical scanning results imaging speed too slow to 3D SRM imaging, traditional MSIM needs 224 frames raw multifocal images to reconstruct a 2D SRM slice, and thus it requires at least 1s to obtain a MSIM optical section image. Therefore, the combination of multiplane prism and MSIM is needed to acquire the 3D information of a certain axial distance through a single shot. Employing a prism with special structure to split the imaging channel into eight channels with different optical path difference, the MSIM system can increase the 3D SRM imaging speed up to eight times compared to that of conventional one. Real-time and fast living SRM volumetric imaging of most thin cell samples are allow to achieve through this fast multifocal structured illumination microscopy based on the multiplane prism (MPP-MSIM). Compared to another fast 3D SRM methods such as fast-SIM based on distorted grating and fast-MSIM based on double-helix phase mask, multiplane prism has almost no chromatic aberration within the spectral range of 500-700 nanometers as well as no limitation in phase period repetition caused by rotational symmetry of the spiral phase, MPP-MSIM is able to reconstruct the real 3D imaging information of thick biological samples with different emission wavelengths, which makes MPP-MSIM a significant development value in fast 3D SRM imaging of biomedical domain