Fast live simultaneous multiwavelength four-dimensional optical microscopy
Fast live simultaneous multiwavelength four-dimensional optical microscopy
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DOI：
10.1073/pnas.1004037107
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发表时间：
2010-09-14
影响因子：
11.1
通讯作者：
Sedat, John W.
中科院分区：
文献类型：
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作者：
Carlton, Peter M.;Boulanger, Jerome;Sedat, John W.
活荧光显微镜具有独特的能力，以探测动态过程，连接分子成分和它们的定位与功能。显微镜的一个关键目标是提高空间和时间分辨率，同时允许识别多个特定成分。我们展示了一种新的显微镜平台，OMX，使亚秒，100万四维数据采集，也提供了访问子衍射结构照明成像。使用该平台以每秒一个3D图像堆栈的速度在完整的酵母细胞周期期间对染色体运动进行成像，揭示了含荧光团细胞的出乎意料的光敏性程度。为了避免细胞分裂的干扰，激发水平必须在低于通常用于成像的水平的100到10，000倍之间衰减。我们表明，图像去噪算法，利用冗余的图像序列在空间和时间上允许恢复的生物信息从低亮度噪声图像，同时保持完整的细胞活力，没有衰落。
Live fluorescence microscopy has the unique capability to probe dynamic processes, linking molecular components and their localization with function. A key goal of microscopy is to increase spatial and temporal resolution while simultaneously permitting identification of multiple specific components. We demonstrate a new microscope platform, OMX, that enables subsecond, multicolor four-dimensional data acquisition and also provides access to sub-diffraction structured illumination imaging. Using this platform to image chromosome movement during a complete yeast cell cycle at one 3D image stack per second reveals an unexpected degree of photosensitivity of fluorophore-containing cells. To avoid perturbation of cell division, excitation levels had to be attenuated between 100 and 10,000x below the level normally used for imaging. We show that an image denoising algorithm that exploits redundancy in the image sequence over space and time allows recovery of biological information from the low light level noisy images while maintaining full cell viability with no fading.