On April 11th 2018, international authoritative magazine Nature Biotechnology (SCI IF: 41.67) published a paper named Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy. The corresponding authors are Prof. Shan Tan from School of Artificial Intelligence and Automation, Huazhong University of Science and Technology and Prof. Liangyi Chen from Institute of Molecular Medicine, Peking University, and the first authors are doctoral student Junchao Fan from HUST, doctoral students Xiaoshuai Huang and Liuju Li from Peking University.
Shan Tan’s group and Liangyi Chen’s group achieve important breakthroughs in super-resolution fluorescence imaging with structured illumination microscopy (SIM) and realize fast, long-term and super-resolution imaging of living cells at a low signal-to-noise ratio.
Due to the existence of diffraction limit, the spatial resolution of the traditional fluorescence micro-imaging technique cannot exceed 200 nanometers. In recent years, especially marked by the Nobel Prize in Chemistry in 2014, super-resolution fluorescence microscopy technology attains a booming development. It enables biologists to observe living cells’ structures and dynamic processes at a more micro-level (in a higher spatial resolution).
San Tan’s group and Liangyi Chen’s group started to cooperate in SIM technology since 2014. After years of efforts, they eventually developed fast, long-term and super-resolution imaging with Hessian SIM. It can not only realize ultra-high-resolution imaging but also realize a long time ultra-high-speed imaging of living cells for the first time. This technology is the ultra-high-resolution fluorescence microscopic imaging technology with the longest imaging time and the highest time resolution currently.
Compared with traditional SIM, Hessian SIM has great breakthroughs in both software and hardware. In hardware, through high numerical aperture objective lens, self-designed polarization rotating glass array and High precision timing control program, Hessian SIM increases the collection ability of photons, shortens the sampling time and then realizes high-speed sampling of images. In software, Hessian SIM designs a new structured light parameter calculation method so that the reconstruction parameters could still be calculated correctly at a low signal-to-noise ratio. Hessian SIM puts forward the Hessian Regular based on multidimensional space continuity for the first time, which realized high-quality super-resolution reconstruction by utilizing the characteristics (continuous in space but not in noise) of cell structure. Hessian SIM also summarized and analyzed the causes of artifacts in traditional SIM imaging, and proposed a set of high-quality reconstruction schemes.
The spatial resolution of Hessian SIM can reach 85 nanometers (about 1/600 to 1/800 of a single hair). When 188 images are taken per second at a high speed, the required illumination is only 8 W/cm2. For its low photobleaching and low phototoxicity, Hessian SIM can keep imaging for 10 minutes and yield 180,000 super-high-resolution images under high-speed sampling in 100 Hz, or keep super-high-resolution imaging for 1 hour without photobleaching under sampling in 1 Hz.
With Hessian SIM, Liangyi Chen’s group analyzes the fusion and recombination process of mitochondria in living cells for the first time. And in the process of observing the fusion and release of neurotransmitter and hormone by intracellular vesicle and plasma membrane, the process of membrane channel formation is visualized.
Junchao Fan and Haosen Liu from Shan Tan’s group were responsible for the design and realization of Hessian SIM reconstruction algorithm, Xiaoshuai Huang and Liuju Li from Liangyi Chen’s group were responsible for the design and construction of hardware platform and related biological experiments. Academician Heping Cheng from Institute of Molecular Medicine, Peking University provided some guiding opinions. The related software and hardware technologies of Hessian SIM have been applied for patent protection in HUST and Peking University, respectively.
