Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy
Fast, long-term, super-resolution imaging with Hessian structured illumination microscopy
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使用 Hessian 结构照明显微镜进行快速、长期、超分辨率成像
DOI：
10.1038/nbt.4115
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发表时间：
2018-05-01
影响因子：
46.9
通讯作者：
Chen, Liangyi
中科院分区：
文献类型：
--
作者：
Huang, Xiaoshuai;Fan, Junchao;Chen, Liangyi
为了提高超分辨率显微镜的时间分辨率和最大成像时间，提出了一种基于Hessian矩阵的结构照明显微镜解卷积算法（Hessian-SIM）。它使用生物结构在多个维度上的连续性作为先验知识来指导图像重建，并获得伪影最小化的SR图像，其光子剂量小于传统SIM使用的10%，同时在低信号强度下大大优于当前算法。Hessian-SIM能够快速成像内质网中的移动囊泡或环，而没有运动伪影，时空分辨率为88 nm和188 Hz。它的高灵敏度允许使用亚毫秒的激发脉冲，然后是暗恢复时间，以减少荧光蛋白的光漂白，从而实现活细胞中肌动蛋白丝的小时延时SR成像。最后，我们观察到的线粒体嵴和结构，据我们所知，以前没有观察到的结构动力学，如囊泡胞吐过程中扩大的融合孔。
To increase the temporal resolution and maximal imaging time of super-resolution (SR) microscopy, 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. Hessian-SIM enables rapid imaging of moving vesicles or loops in the endoplasmic reticulum without motion artifacts and with a spatiotemporal resolution of 88 nm and 188 Hz. 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 of actin filaments in live cells. Finally, we observed the structural dynamics of mitochondrial cristae and structures that, to our knowledge, have not been observed previously, such as enlarged fusion pores during vesicle exocytosis.