Validation of peptide epitope microarray experiments and extraction of quality data
Validation of peptide epitope microarray experiments and extraction of quality data
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DOI：
10.1016/j.jim.2007.07.015
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发表时间：
2007-12-01
影响因子：
2.2
通讯作者：
Reilly, Marie
中科院分区：
文献类型：
--
作者：
Nahtman, Tatjana;Jernberg, Alexander;Reilly, Marie
简介：在过去的十年中，抗原微阵列载玻片的发展使得能够同时测量血清对单个生物样品中数百种肽的反应性。尽管取得了相当大的科学进步，但在这些幻灯片产生的数据的质量、分析和解释方面仍然存在许多问题。目前还没有公认的方法来指导数据分析，研究人员使用各种各样的统计方法和软件工具。我们设计并实施了一个实验室实验来评估肽微阵列数据的可靠性和测量范围，并提出了用于预处理的图形和统计程序，以便可以提取高质量的数据来解决生物学假设。跨越蛋白质Ag85A、Ag85B、CFP10、MPT51/MPB51、选择TB 10.4和ESAT-6作为筛选结核分枝杆菌（MTB）血清反应性的范例。我们探索了各种定量和图形方法来展示幻灯片中的响应。我们对来自5个TB阳性个体的样本进行了重复测定，以检查重复性，并使用线性混合模型来研究变异性的各种来源，并评估测量范围。我们使用我们的方法来提取数据从5 TB阳性的个人和5个健康对照，并分析瓷砖"归一化"响应：使用免费提供的SAM package.Results：前景与背景信号（油的对数尺度）的比率提供了一个适当的响应指数。一个二维图形显示清晰地说明了幻灯片上的对照和肽特征的反应。对复制载玻片的混合模型分析发现，操作员、天数和实验之间的瓦片测定具有较高的重现性。测量范围也令人满意。我们的分析表明，从5 TB阳性患者和5个健康对照组的标准化反应的363 pep：ides评估有显着较高的反应TB阳性group.Conclusions：精心设计的实验室实验和严格的统计分析，可以使去除技术文物产生高质量的肽阵列数据，解决生物学假设。这些仪器能够在载玻片和/或载玻片批次之间进行有效的比较，将为未来针对广泛的B细胞表位进行高含量血清反应性分析的比较分析保驾护航。(C)2007 Elsevier B.V.保留所有权利。
Introduction: Within the last decade, the development of antigen microarray slides has enabled the simultaneous measurement of serum reactivity to hundreds of peptides in a single biological sample. Despite this considerable scientific progress, many issues remain regarding the quality, analysis and interpretation of the data these slides produce. There is currently no accepted approach to guide data analysis, and researchers use a wide variety of statistical methods and software tools. We designed and implemented a laboratory experiment to assess the reliability and range of measurement of peptide microarray data, and present graphical and statistical procedures for pre-processing so that quality data can be extracted for addressing biological hypotheses.Methods: Synthetic peptides spanning the proteins Ag85A, Ag85B, CFP10, MPT51/MPB51, TB10.4 and ESAT-6 were chosen as a paradigm to screen for serum reactivity to Mycobacteria tuberculosis (MTB). We explored various quantitative and graphical methods for presenting the responses from a slide. We replicated assays of samples from five TB-positive individuals to examine reproducibility, and used linear mixed models to investigate the various sources of variability, and to assess the range of measurement. We use our methods to extract data from the five TB-positive individuals and five healthy controls, and analyse tile "normalized" response:; using the freely available SAM package.Results: The ratio of foreground to background signal (oil a log scale) provides an appropriate response index. A two-dimensional graphical display cleary illustrates the responses from the control and peptide features on a slide. Mixed model analysis of the replicated slides found a high reproducibility of tile assay between operators, days and experiments. The range of measurement was also satisfactory. Our analysis of the normalized responses from the five TB-positive patients and five healthy controls suggested that 10 of the 363 pep:ides assessed had significantly higher responses in the TB-positive group.Conclusions: Carefully designed laboratory experiments and rigorous statistical analysis can enable the removal of technical artefacts to produce quality peptide array data for addressing biological hypotheses. These instruments, which enable valid comparisons across slides and/or batches of slides, will escort future comparative analyses targeting high content serum reactivity profiling against a broad array of B-cell epitopes. (C) 2007 Elsevier B.V. All rights reserved.