Pellis Linette, Franssen-van Hal Nicole L W, Burema Jan, Keijer Jaap
Food Bioactives Group, RIKILT-Institute of Food Safety, 6700 AE Wageningen, The Netherlands.
Physiol Genomics. 2003 Dec 16;16(1):99-106. doi: 10.1152/physiolgenomics.00111.2003.
We show that the intraclass correlation coefficient (ICC) can be used as a relatively simple statistical measure to assess methodological and biological variation in DNA microarray analysis. The ICC is a measure that determines the reproducibility of a variable, which can easily be calculated from an ANOVA table. It is based on the assessment of both systematic deviation and random variation, and it facilitates comparison of multiple samples at once. We used the ICC first to optimize our microarray data normalization method and found that the use of median values instead of mean values improves data correction. Then the reproducibility of different labeling methods was evaluated, and labeling by indirect fluorescent dye incorporation appeared to be more reproducible than direct labeling. Finally, we determined optimal biopsy sampling by analyzing overall variation in gene expression. The variation in gene expression of rectal biopsies within persons decreased when two biopsies were taken instead of one, but it did not considerably improve when more than two biopsies were taken from one person, indicating that it is sufficient to use two biopsies per person for DNA microarray analysis under our experimental conditions. To optimize the accuracy of the microarray data, biopsies from at least six different persons should be used per group.
我们表明，组内相关系数（ICC）可作为一种相对简单的统计量度，用于评估DNA微阵列分析中的方法学和生物学变异。ICC是一种确定变量可重复性的量度，可轻松从方差分析表中计算得出。它基于对系统偏差和随机变异的评估，便于同时比较多个样本。我们首先使用ICC来优化微阵列数据归一化方法，发现使用中位数而非平均值可改善数据校正。然后评估了不同标记方法的可重复性，结果显示间接荧光染料掺入标记似乎比直接标记更具可重复性。最后，我们通过分析基因表达的总体变异来确定最佳活检采样。当采集两份而非一份直肠活检样本时，个体内直肠活检样本的基因表达变异会降低，但当从一个人身上采集超过两份活检样本时，变异并未显著改善，这表明在我们的实验条件下，每人使用两份活检样本进行DNA微阵列分析就足够了。为了优化微阵列数据的准确性，每组应使用至少六个不同个体的活检样本。
