High spatiotemporal resolution fluorescence imaging of biological samples in vivo
陈良怡
北京大学
摘要: Here we will present three pieces of high-resolution fluorescence microscopy methods we invented for live sample imaging. The first one is for in vivo imaging, which is a series of fast, high-resolution, miniaturized two-photon microscope (FHIRM-TPM), which can be used to resolve single spine in freely-behaving animals and achieved volumetric or mulitplane imaging over an axial distance of 180 µm with an interplane switch time of less than 1.5 ms. Further, we engineered the headpiece for repeated mounting and dismounting, and demonstrated its robustness by recording neuronal activities from the same brain region over a time frame of several weeks.
The second method is for live cell long-term super-resolution (SR) imaging. We have developed a deconvolution algorithm for structured illumination microscopy based on Hessian matrixes (Hessian-SIM). It uses the continuity of biological structures in multiple dimensions as a priori knowledge to guide image reconstruction and attains artifact-minimized SR images with less than 10% of the photon dose used by conventional SIM while substantially outperforming current algorithms at low signal intensities. Its high sensitivity allows the use of sub-millisecond excitation pulses followed by dark recovery times to reduce photobleaching of fluorescent proteins, enabling hour-long time-lapse SR imaging in live cells.
The third technology is a dual-mode SR microscopy for highlighting molecules as well as a holistic view of related interacting organelles in live cells. It is a combination of two-dimensional Hessian-SIM with label-free three-dimensional optical diffraction tomography (ODT), term SR fluorescence-assisted diffraction computational tomography (SR-FACT). The ODT module is capable of resolving mitochondria, lipid droplets, the nuclear membrane, chromosomes, the tubular endoplasmic reticulum and lysosomes. These works demonstrate the unique capabilities of SR-FACT, which suggest its wide applicability in cell biology in general.
个人简介: 北京大学未来技术学院博雅特聘教授,博士,博士生导师,发明了一系列的高时空分辨率生物医学成像手段,获得国家自然科学基金委医学部杰出青年基金、重大研究计划集成项目等多项资助。不仅推动了国内外许多合作者的生物医学基础研究工作,还将原创技术转化成为国内急需的高端显微镜产品,解决国内高端显微镜产品被国外厂商“卡脖子”的现状。主要发明包括:高分辨率微型化双光子显微镜,开创高分辨率在体成像领域;超灵敏海森结构光超分辨率显微镜,实现通用的活细胞超分辨率成像;活细胞荧光-无标记双模态超分辨率显微镜,揭示活细胞细胞器互作组以及找到新细胞器;提出活细胞超分辨率病理学的概念,并揭示佩梅病临床疾病表型机制以及筛选精准对症药物。工作曾入选了“2017年中国科学十大进展”,“2017年中国十大医学科技新闻”,“2017年中国生命科学十大进展”,“2018年中国光学十大进展”等奖项。