Three-Dimension Resolution Enhanced Microscopy Based on Parallel Detection

Pixel reassignment image scanning microscopy (PRISM) is a useful tool to improve the resolution of confocal laser scanning microscopy (CLSM) only equipped with a detector array. However, while it can improve the lateral resolution, it has little effect on the axial resolution. Here, new microscopy has been proposed which combines three-dimension fluorescence emission difference microscopy (3D FED) with PRISM to further improve three-dimension resolution. We call this method three-dimension pixel reassignment fluorescence emission difference microscopy (3D-PRFED). Detailed theoretical analysis and simulation are presented in this paper. Additionally, the performance of lateral and axial resolution improvement of this method has been demonstrated by imaging 100 nm fluorescent beads and nuclear pore complexes samples. Experiment results show that this method in our system can improve lateral resolution by a factor of 1.85 and axial resolution by a factor of 1.48 compared with CLSM.