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PKU Researchers Developed Large-Field High-Resolution Two-Photon Digital Scanned Light-Sheet Microscopy

2014-12-26 11:07:00


A research team from Peking University (PKU)’s Institute of Molecular Medicine has developed a novel two-photon three-axis digital scanned light-sheet microscope (2P3A-DSLM). To achieve high axial resolution (thin light sheet) and large field of view simultaneously, it employs a technology based on ultrafast axial scanning of illumination focal spot with a tunable acoustic gradient (TAG) index device. 2P3A-DSLM is able to achieve minimal photobleaching, high axial and temporal resolution with deep-tissue imaging. 2P3A-DSLM demonstrates its superior performance by resolving subcellular structures and tracking single-mitochondrion dynamic processes in live model organisms.
 
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This new technique provides a sub-micron axial resolution, a large field of view of 170 × 170 μm2 in deep tissues, and a high temporal resolution limited only by the detection camera. In vivo imaging with 2P3A-DSLM resolved subcellular structures and dynamic processes in small organisms.A TAG lens uses acoustic wave to radially excite a fluid-filled cylindrical cavity and produce continuous changes in refractive power that enables rapid change of its axial focal plane within 10 μs.

Due to the small size of chromosomes, resolving chromosomal segregation during cell division in cultured single cells or at the earlier stage of embryo development usually requires high-NA, high-magnification objectives. Using 2P3A-DSLM, it is able to visualize hundreds of nuclei in the heart of a day 3 zebrafish simultaneously. Chromosome structures from a few dividing cells could be well resolved due to the extremely thin light sheet, low background and high contrast provided by the new imaging method.

The first author is Zong Weijian, a PhD candidate from the Institute of Molecular Medicine, PKU and the Academy of Military Science. Professor Liangyi Chen from the Institute of Molecular Medicine and Sun Yujjie from Biodynamics Optical Imaging Center are corresponding authors.  

By Lewis Won Cheng Yi