Super-resolution fluorescence imaging is one of the most important breakthroughs in microscopic optical imaging in this century and was awarded the 2014 Nobel Prize in Chemistry. Among them, super-resolution positioning imaging is a representative super-resolution fluorescence imaging technology. Based on Total Internal Reflection Fluorescence Microscopy (TIRF), the technology combines single-molecule imaging with high-precision molecular localization algorithms to achieve ultra-high spatial resolution of 20-30 nm, providing an unprecedented opportunity for biomedical research . However, the conventional super-resolution positioning imaging system, due to the single molecule imaging using the weak light detector EMCCD, has a limited imaging field of view of mostly 30 μm to 30 μm. In addition, the spatial resolution of conventional super resolving positioning imaging systems is generally not uniform due to the single-molecule fluorescence excitation using a Gaussian spot: the resolution of the field edge is typically much lower than the resolution of the field center. Therefore, how to increase the imaging field of view and achieve large field of view super-resolution positioning imaging while ensuring uniform imaging resolution is a hot spot in recent years.
Under the guidance of Prof. Huang Zhenli from Britton Chance Biomedical Photonics Research Center of Wuhan National Laboratory for Optoelectronics, a 2013 doctoral student Zhao Zeyu proposed a high-power uniform illumination scheme suitable for large-field super-resolution positioning imaging system, and realized Large-field super-resolution positioning imaging system based on epi-illumination. The system provides uniform spatial resolution (40 nm) with an imaging field of up to 221 μm Å and 221 μm, which is more than 50 times that of conventional super-resolution positioning imaging fields and is the largest imaging field reported in the literature About 5 times. In addition to meeting the high power requirements of large-field imaging, this scheme also outputs a square homogenization spot to match the square detector shape in low-light cameras. June 2, 2017 This research is published in Optics Express, Vol. 25, Iss. 12, pp..
Fig. Illumination spot uniformity (ab) and large-field super-resolution localization imaging results (cd) for COS-7 cells. The scale in c is 50mm
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