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首页> 外文期刊>Cell and Tissue Research >The border-to-border distribution method for analysis of cytoplasmic particles and organelles
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The border-to-border distribution method for analysis of cytoplasmic particles and organelles

机译：用于细胞质颗粒和细胞器分析的边界到边界分布方法

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摘要

Comparing the distribution of cytoplasmic particles and organelles between different experimental conditions can be challenging due to the heterogeneous nature of cell morphologies. The border-to-border distribution method was created to enable the quantitative analysis of fluorescently labeled cytoplasmic particles and organelles of multiple cells from images obtained by confocal microscopy. The method consists of four steps: (1) imaging of fluorescently labeled cells, (2) division of the image of the cytoplasm into radial segments, (3) selection of segments of interest, and (4) population analysis of fluorescence intensities at the pixel level either as a function of distance along the selected radial segments or as a function of angle around an annulus. The method was validated using the well-characterized effect of brefeldin A (BFA) on the distribution of the vesicular stomatitis virus G protein, in which intensely labeled Golgi membranes are redistributed within the cytoplasm. Surprisingly, in untreated cells, the distribution of fluorescence in Golgi membrane-containing radial segments was similar to the distribution of fluorescence in other G protein-containing segments, indicating that the presence of Golgi membranes did not shift the distribution of G protein towards the nucleus compared to the distribution of G protein in other regions of the cell. Treatment with BFA caused only a slight shift in the distribution of the brightest G protein-containing segments which had a distribution similar to that in untreated cells. Instead, the major effect of BFA was to alter the annular distribution of G protein in the perinuclear region.
机译：由于细胞形态的异质性，比较不同实验条件之间的细胞质颗粒和细胞器的分布可能具有挑战性。创建了边界到边界的分配方法，以便能够通过共聚焦显微镜获得的图像对荧光标记的细胞质颗粒和多个细胞的细胞器进行定量分析。该方法包括四个步骤：（1）荧光标记细胞的成像;（2）将细胞质的图像划分为放射状片段;（3）选择感兴趣的片段;（4）荧光强度的群体分析。像素级别取决于沿所选径向线段的距离或围绕圆环的角度的函数。使用布雷菲德菌素A（BFA）对水泡性口炎病毒G蛋白分布的良好表征作用验证了该方法，其中强烈标记的高尔基体膜在细胞质内重新分布。令人惊讶的是，在未经处理的细胞中，含高尔基膜的径向片段中的荧光分布与其他含G蛋白的片段中的荧光分布相似，这表明高尔基膜的存在不会使G蛋白的分布向核移动与G蛋白在细胞其他区域的分布相比。用BFA处理只会导致最亮的含G蛋白片段的分布发生轻微变化，该分布与未经处理的细胞相似。取而代之的是，BFA的主要作用是改变了核周区域G蛋白的环状分布。

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