Bergers E, Montironi R, van Diest P J, Prete E, Baak J P
Department of Pathology, Free University Hospital, Amsterdam, The Netherlands.
Hum Pathol. 1996 Jun;27(6):553-60. doi: 10.1016/s0046-8177(96)90161-6.
Conflicting prognostic results have been published as to the DNA variables, such as DNA ploidy, DNA index, and % S-phase cells for breast cancer patients. These variables can be obtained by interpreting DNA histograms by cell cycle analysis. Explanations for these conflicting results might be found on the level of the interpretation of the DNA histograms. In a previous study, the semi automated cell cycle analysis computer program MultiCycle (Phoenix Flow Systems, San Diego, CA) showed high intralaboratory reproducibility. However, what types of DNA histograms may cause disagreements was still unclear. The aim of this study was to determine the interlaboratory reproducibility of MultiCycle-based cell cycle analysis of 1,295 flow cytometric DNA histograms derived from fresh frozen breast cancer material and to clarify potential sources of interobserver variation when analyzing DNA histograms. DNA ploidy classification into diploid, hyperdiploid, tetraploid, hypertetraploid, and multiploid showed an interlaboratory agreement of 94% (kappa value = 0.92). The 6% discrepancies (n = 74) were caused by tetraploid peaks, as established in one laboratory, which shifted outside the tetraploid region on reanalysis by the other laboratory (37%), shoulders sometimes interpreted as peaks (24%), small peaks not always recognized as such (24%), fitting failures (10%), and overlooking of tetraploid peaks (5%). Furthermore, the cell cycle analysis variables showed variable reproducibility. The % S-phase cells of the first, second, and third cell cycle showed overall a moderate reproducibility (0.62 < or = R < or = 0.79), but the average % S-phase cells and the average aneuploid % S-phase cells were more reproducible with correlation coefficients of 0.89 and 0.81, respectively. The coefficient of variation of the G0/G1 peak of the first cell cycle, the DNA indices and the % diploid cells were highly reproducible (R > or = 0.94), and the % G2/M-phase cells of the first, second, and third cell cycle were poorly reproducible (0.22 < or = R < or = 0.68). When a cut-point was used at the mean value of 7% for the average % S-phase cells, the number of "threshold discrepancy cases" was 6%. Sources of variation for cell cycle analysis were variations in the debris correction procedures, disagreement about the modes of the aneuploid peaks, disagreement about small peaks, shoulders sometimes interpreted as peaks, and overlooking of tetraploid peaks.
关于乳腺癌患者的DNA变量，如DNA倍体、DNA指数和S期细胞百分比，已经发表了相互矛盾的预后结果。这些变量可以通过细胞周期分析解释DNA直方图来获得。对于这些相互矛盾的结果的解释可能可以在DNA直方图的解释层面找到。在之前的一项研究中，半自动细胞周期分析计算机程序MultiCycle（Phoenix Flow Systems，圣地亚哥，加利福尼亚州）显示出较高的实验室内重复性。然而，何种类型的DNA直方图可能导致分歧仍不清楚。本研究的目的是确定基于MultiCycle的对1295份来自新鲜冷冻乳腺癌材料的流式细胞术DNA直方图进行细胞周期分析的实验室间重复性，并在分析DNA直方图时阐明观察者间差异的潜在来源。DNA倍体分类为二倍体、超二倍体、四倍体、超四倍体和多倍体，显示实验室间一致性为94%（kappa值=0.92）。6%的差异（n = 74）是由四倍体峰引起的，如在一个实验室中确定的，在另一个实验室重新分析时，这些峰移出了四倍体区域（37%），有时将肩部解释为峰（24%），小峰并非总是被识别为小峰（24%），拟合失败（10%），以及忽略四倍体峰（5%）。此外，细胞周期分析变量显示出不同的重复性。第一、第二和第三个细胞周期的S期细胞百分比总体显示出中等重复性（0.62≤R≤0.79），但平均S期细胞百分比和平均非整倍体S期细胞百分比的重复性更高，相关系数分别为0.89和0.81。第一个细胞周期的G0/G1峰、DNA指数和二倍体细胞百分比的变异系数具有高度重复性（R≥），而第一、第二和第三个细胞周期的G2/M期细胞百分比的重复性较差（0.22≤R≤0.68）。当将平均S期细胞百分比的切点设定为7%的平均值时，“阈值差异病例”的数量为6%。细胞周期分析的变异来源包括碎片校正程序的差异、关于非整倍体峰模式的分歧、关于小峰的分歧、有时将肩部解释为峰以及忽略四倍体峰。
