Author Affiliations
Abstract
1 Southern University of Science and Technology, College of Engineering, UTS-SUSTech Joint Research Centre for Biomedical Materials and Devices, Department of Biomedical Engineering, Shenzhen, China
2 City University of Hong Kong, Department of Biomedical Engineering, Hong Kong, China
3 Peking University, College of Future Technology, Department of Biomedical Engineering, Beijing, China
4 University of Technology Sydney, Institute for Biomedical Materials and Devices (IBMD), Faculty of Science, Sydney, Australia
In light-sheet fluorescence microscopy, the axial resolution and field of view are mutually constrained. Axially swept light-sheet microscopy (ASLM) can decouple the trade-off, but the confocal detection scheme using a rolling shutter also rejects fluorescence signals from the specimen in the field of interest, which sacrifices the photon efficiency. Here, we report a laterally swept light-sheet microscopy (LSLM) scheme in which the focused beam is first scanned along the axial direction and subsequently laterally swept with the rolling shutter. We show that LSLM can obtain a higher photon efficiency when similar axial resolution and field of view can be achieved. Moreover, based on the principle of image scanning microscopy, applying the pixel reassignment to the LSLM images, hereby named iLSLM, improves the optical sectioning. Both simulation and experimental results demonstrate the higher photon efficiency with similar axial resolution and optical sectioning. Our proposed scheme is suitable for volumetric imaging of specimens that are susceptible to photobleaching or phototoxicity.
light-sheet fluorescence microscopy image scanning microscopy volumetric imaging pixel reassignment Advanced Photonics Nexus
2023, 2(1): 016001
Author Affiliations
Abstract
1 State Key Laboratory of Modern Optical Instrumentation College of Optical Science and Engineering, Zhejiang University Hangzhou, Zhejiang 310027, P. R. China
2 Ningbo Research Institute, Zhejiang University Ningbo 315100, P. R. China
3 Collaborative Innovation Center of Extreme Optics, Shanxi University Taiyuan, Shanxi 030006, P. R. China
4 College of Electronics and Information Engineering Shanghai University of Electric Power, Shanghai 200090, P. R. China
5 Key Laboratory of Optoelectronic Science and Technology for Medicine Ministry of Education and Fujian Provincial Key Laboratory for Photonics Technology Fujian Normal University, Fuzhou 350007, P. R. China
Image scanning microscopy based on pixel reassignment can improve the confocal resolution limit without losing the image signal-to-noise ratio (SNR) greatly [C. J. R. Sheppard, "Super-resolution in confocal imaging," Optik 80(2) 53–54 (1988). C. B. Müller, E. Jorg, "Image scanning microscopy, "Phys. Rev. Lett. 104(19) 198101 (2010). C. J. R. Sheppard, S. B. Mehta, R. Heintzmann, "Superresolution by image scanning microscopy using pixel reassignment," Opt. Lett. 38(15) 2889–2892 (2013)]. Here, we use a tailor-made optical fiber and 19 avalanche photodiodes (APDs) as parallel detectors to upgrade our existing confocal microscopy, termed as parallel-detection super-resolution (PDSR) microscopy. In order to obtain the correct shift value, we use the normalized 2D cross correlation to calculate the shifting value of each image. We characterized our system performance by imaging fluorescence beads and applied this system to observing the 3D structure of biological specimen.
Pixel reassignment SIM parallel-detection super-resolution (PDSR) microsc normalized cross-correlation algorithm Journal of Innovative Optical Health Sciences
2019, 12(6):
