Adaptive correction technique for 3D reconstruction of fluorescence microscopy images
Adaptive correction technique for 3D reconstruction of fluorescence microscopy images
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DOI：
10.1002/jemt.20536
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发表时间：
2008-02-01
影响因子：
2.5
通讯作者：
Opas, M.
中科院分区：
文献类型：
--
作者：
Guan, Y. Q.;Cai, Y. Y.;Opas, M.
高分辨率成像的最新进展为体外和体内细胞和组织内的结构关系提供了有价值的新见解。这种分析通常通过使用共聚焦或去卷积显微镜的光学切片来进行。然而，3D图像的重建由于所使用的介质的有限透明度而随着深度的增加而遭受光散射和吸收。荧光染料的光漂白特别麻烦，通常在光学切片期间对信号损失的唯一补救是减少切片的数量。这导致体素的x-y和z维度的差异，这导致图像的原始堆叠的垂直失真并且需要插值。插值是必要的，以填补在原始图像堆栈中的连续部分之间的间隙，以获得立方体素。本手稿描述了一种新的方法，用于自适应补偿的光强度衰减的荧光显微镜图像的堆栈是基于光衰减的物理模型。首先，我们使用一种快速插值技术来生成一个基于立方体体素的体积堆栈的帮助下的贡献查找表。利用贡献查找表，避免了多次计算，这在不损害恢复过程的准确性的情况下显著地减少了计算时间。第二，处理所得到的体积内的每个部分，以校正由于光漂白、散射和吸收而改变的强度值。该方法允许定义堆栈中的最后一个良好部分，然后自动进行校正。
Recent advances in high-resolution imaging have provided valuable novel insights into structural relationships within cells and tissues both in vitro and in vivo. An analysis of this kind is regularly done by optical sectioning using either confocal or deconvolution microscopy. However, the reconstruction of 3D images suffers from light scattering and absorption with increasing depth by finite transparency of the used media. Photobleaching of fluorochromes has been especially troublesome and often the only remedy for loss of signal during optical sectioning is to reduce the number of sections. This causes disparities in the x-y and z dimensions of voxels, which lead to vertical distortion of the original stack of images and necessitates interpolation. Interpolation is necessary to fill up the gaps between consecutive sections in the original image stack to obtain cubic voxels. The present manuscript describes a novel method for adaptive compensation of attenuation of light intensity in stacks of fluorescence microscopy images that is based on a physical model of light attenuation. First, we use a fast interpolation technique to generate a cubic voxel-based volume stack with the aid of a contribution look up table. With the contribution look up table, multiple calculations are avoided, which substantially reduces the computational time without compromising the accuracy of the restoration procedure. Second, each section within the resulting volume is processed to rectify its intensity values that have been altered due to photobleaching and scattering and absorption. The method allows to define the last good section in the stack and the correction is then done automatically.