Recently, researcher Zheng Wei of Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, in collaboration with Professor Hari Shroff of the National Institutes of Health, successfully developed a new two-photon excited super-resolution optical microscopy system that has both super-resolution optical microscopy And large depth 3D imaging capabilities to advance the depth of optical super-resolution imaging to record-breaking 250 microns, the corresponding research results published in the August 2017 issue of Adaptive optics improves multiphotonsuper-resolution imaging "Nature - Methodology" (Nature - Methods), Zheng Wei researcher is the article's first author and correspondent.
"Fine-looking" and "deep-looking" are the two major challenges in the field of optical microscopy. After decades of unremitting efforts by researchers, both in terms of "seeing the details" and "seeing the depth" A number of innovations have emerged and great successes have been achieved. However, optical microscopy imaging with both the ability to "see the details" and the "see the depth" is rare.
In this study, Zheng et al. Compared Two-Photon Microscopy (TPM) with deep biological tissue imaging capabilities and Instant Structured Light Microscopy (Instant Structured Light Microscopy) with super-resolution imaging , ISIM) organically combine to achieve two-photon excitation of super-resolution microscopic imaging. At the same time, the researchers successfully overcome the problem of wavefront phase distortion caused by biological tissues by using adaptive optics (AO) technology to finally achieve a horizontal resolution of 176 nm, a resolution of 729 nm and a resolution of 250 μm Depth of the imaging effect. Using this technique, high-resolution three-dimensional imaging of cells, nematode embryos and larvae, fruit fly brain slices and zebrafish embryos can be performed. The imaging results are significantly better than the traditional two-photon imaging. It is worth mentioning that, because the technology improves the photon utilization efficiency, thereby reducing the required laser power, the development of nematode embryos can be carried out for a long time, high-definition three-dimensional dynamic observation. It has no effect on nematode embryo development during up to one hour of continuous three-dimensional imaging, and this technique has an important role in embryonic development research.
The research was supported by the projects supported by the National Natural Science Foundation of China, the State Key Basic Research Development (973) Program and the Peacock Project of Shenzhen Overseas High-level Talents Innovation and Entrepreneurship.
The left panel shows the contrast between the results of two-photon imaging (2P WF), two-photon super-resolution imaging (2P ISIM) and two-photon super resolution (2P ISIM AO) combined with adaptive optics. In contrast to the three-dimensional imaging of cells located in the depth of 150 μm of collagen gel, the resolution of the new technology has been significantly improved both in horizontal and vertical directions. The lower right picture shows the three-dimensional observation of the nematode during embryonic development for 1 hour. The normal cell division process proves that the technology can be used for the dynamic research of embryonic development.
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