间期细胞核中的荧光点计数

Hans Netten, Lucas J van Vliet, Hans Vrolijk, Willem C R Sloos, Hans J Tanke, Ian T Young
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引用次数: 56

摘要

荧光原位杂交允许计数间期细胞核中的染色体异常。这个过程叫做点计数。为了估计每个细胞的染色体分布,必须对大量细胞进行分析,特别是当异常细胞的频率较低时。点计数的自动化是可取的,因为手动计数是乏味、疲劳和耗时的。我们开发了一种全自动荧光显微镜系统,可以在间期细胞核中为一个探针计数荧光杂交点。该系统使用两种荧光染料——一种用于DNA杂交点,另一种用于细胞核。图像采集采用了全自动扫描程序。获取图像后,必须对其进行分析,以找到细胞核并检测点。这篇文章的重点是网点检测程序。提出了三种不同的算法。讨论了“重叠”点和分裂点的问题。自动点计数器已经在许多正常标本上进行了测试,其中DAPI用于细胞核计数器染色,着丝粒探针用于标记12号染色体。载玻片含有来自培养血液的淋巴细胞。对不同算法的性能进行了评估,并与手动获得的结果进行了比较。自动计数的结果与手动计数的结果近似。
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Fluorescent dot counting in interphase cell nuclei

Fluorescence in situ hybridization allows the enumeration of chromosomal abnormalities in interphase cell nuclei. This process is called dot counting. To estimate the distribution of chromosomes per cell, a large number of cells have to be analysed, particularly when the frequency of aberrant cells is low. Automation of dot counting is desirable because manual counting is tedious, fatiguing, and time consuming. We have developed a completely automated fluorescence microscope system that counts fluorescent hybridization dots for one probe in interphase cell nuclei. This system works with two fluorescent dyes—one for the DNA hybridization dots and one for the cell nucleus. A fully automated scanning procedure has been used for the image acquisition. After an image is acquired it has to be analysed in order to find the nuclei and to detect the dots. This article focuses upon the dot detection procedure. Three different algorithms are presented. The problems of ‘overlapping’ dots and split dots are discussed. The automated dot counter has been tested on a number of normal specimens where DAPI was used for the nucleus counter stain and a centromeric probe was used to mark the chromosome 12. The slides contained lymphocytes from cultured blood. The performance of the different algorithms has been evaluated and compared with manually obtained results. The automated counting results approximate the results of manual counting.

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