Oud P S, Bauwens A, Nauwelaers F A
Becton Dickinson Cellular Imaging Systems, Leiden, The Netherlands.
Acta Cytol. 1997 Jan-Feb;41(1):188-96. doi: 10.1159/000332322.
To test a dual DNA-nuclear antigen staining method for multiparameter absorption image analysis.
MCF 7 cells, grown on glass slides, served as a model to test the staining technique. For DNA, Feulgen-based CAS quantitative DNA staining, and for nuclear antigen, alkaline phosphatase-based immunocytochemical staining with CAS Red as the chromogen, were used. MIB-1, estrogen and progesterone receptors were used as examples of nuclear antigen staining. Measurements were performed with the DISCOVERY image analyzer.
Scatterplots, in which the nuclear antigen content was plotted against the DNA content, were obtained. Immunostain-positive and -negative populations could be discriminated. These cells were visualized in image galleries. The DNA histograms of the positive and negative cells showed no change in coefficient of variation or integrated optical density ratio of the G0, G1 and G2 + M peaks as compared to single DNA staining. The intensity of the immunostain increased as compared to the single immunostaining result.
This staining technique allows the simultaneous accurate measurement of costained DNA and antigen within the same nucleus. This opens the possibility for studies in which nuclear antigen expression is monitored during the cell cycle or in cells of different ploidy classes. Identified cells can also be visualized by presentation in an image gallery or by relocation on the slide. This can support the analysis of clinical samples, where cytometric data can be correlated with and confirmed by visual diagnosis.
测试一种用于多参数吸收图像分析的双DNA-核抗原染色方法。
在载玻片上培养的MCF 7细胞用作测试染色技术的模型。对于DNA，采用基于福尔根的CAS定量DNA染色；对于核抗原，采用以碱性磷酸酶为基础、以CAS Red为显色剂的免疫细胞化学染色。以MIB-1、雌激素和孕激素受体作为核抗原染色的示例。使用DISCOVERY图像分析仪进行测量。
获得了散点图，其中核抗原含量与DNA含量相对应绘制。免疫染色阳性和阴性群体能够被区分。这些细胞在图像库中可视化。与单一DNA染色相比，阳性和阴性细胞的DNA直方图在G0、G1和G2 + M峰的变异系数或积分光密度比方面没有变化。与单一免疫染色结果相比，免疫染色强度增加。
这种染色技术能够在同一细胞核内同时准确测量共染色的DNA和抗原。这为在细胞周期或不同倍体类别的细胞中监测核抗原表达的研究开辟了可能性。通过在图像库中展示或在载玻片上重新定位，也能够使识别出的细胞可视化。这可以支持临床样本的分析，在临床样本分析中，细胞计数数据可以与视觉诊断相关联并通过视觉诊断得到证实。
