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目的 :研究彩色颗粒分析软件 (colourpelletamalysissystem ,CPAS)在定量视网膜神经节细胞方面的准确性、可靠性及可重复性。方法 :14张正常大鼠荧光金逆行标记的视网膜神经节细胞照片 ,放大倍数为 12 5倍 ,通过扫描仪扫入计算机 ,使用彩色颗粒分析软件对每张照片的节细胞数进行定量分析。采用计算机自动设定阈值的方法 ,根据灰度对节细胞进行着色 ,再用其自动切分功能对每个节细胞的边界进行界定。删除直径 3μm以下干扰亮点。对自动切分遗漏的节细胞进行人工切分。本研究选用 2名操作员分别进行节细胞的人工计数和计算机计数。观察 2名操作员的节细胞的人工计数之间、节细胞的计算机计数之间是否存在显著性差异 ,比较同一操作员的 2次节细胞计算机计数之间是否存在显著性差异。结果 :2名操作人员的人工计数之间、计算机计数之间均无显著性差异 (两者均 p >0 0 5 )、同一操作员 2次计算机计数之间无显著性差异(p >0 0 5 )。结论 :彩色颗粒分析软件对荧光标记的视网膜节细胞定量分析准确、可靠 ,可重复性好 ,该方法简单易学 ,操作容易。但在定量分析过程中需要对画面进行细致的人工观察 ,部分节细胞需要人工圈定。
Objective: To study the accuracy, reliability and repeatability of colourpelletamalysissystem (CPAS) in quantitative retinal ganglion cells. Methods: Fourteen normal rat retinal ganglion cells retrogradely labeled with fluorescent gold were enrolled in this study. The magnification was 12 5 times, which was swept into the computer by scanner. Quantitative analysis of the number of cells in each photograph was performed by color particle analysis software. The computer automatically set the threshold method, according to the grayscale of the ganglion cells, and then use its automatic segmentation function of each section of cell boundaries. Remove the following interference diameter 3μm bright spots. Artificial segmentation of missing section cells. In this study, two operators were selected for manual counting of ganglion cells and computer counting. Observe whether there is a significant difference between the manual counts of ganglion cells and the computer counts of ganglion cells between the two operators and whether there is a significant difference between the two computers of the same operator in terms of computer counts. Results: There was no significant difference between the two groups of operators (P> 0.05). There was no significant difference between the two operators (p> 0 0) 5). Conclusion: The color particle analysis software has the advantages of accurate, reliable and reproducible fluorescence labeled retinal ganglion cells. The method is easy to learn and easy to operate. However, in the process of quantitative analysis, detailed manual observation of the picture is required, and some sections of cells need manual delineation.