Validation of a novel, fully integrated and flexible microarray benchtop facility for gene expression profiling -: art. no. e151
Validation of a novel, fully integrated and flexible microarray benchtop facility for gene expression profiling -: art. no. e151
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DOI：
10.1093/nar/gng151
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发表时间：
2003-12-01
影响因子：
14.9
通讯作者：
St채hler, PF
中科院分区：
文献类型：
--
作者：
Baum, M;Bielau, S;St채hler, PF
在这里，我们描述了一种新的微阵列平台，它集成了在研究人员的实验室工作台上的紧凑型仪器中执行任何基于阵列的实验所需的所有功能。在三维微流控反应载体的通道内，通过光激活工艺原位合成了寡核苷酸探针。阵列可根据用户要求在数小时内设计和生产。它们是在全自动工作流程中处理的。我们对这个新平台的动态范围、鉴别能力、生物结果的可重复性和准确性进行了描述。该仪器以1:10 000的频率检测样品rna。检测是定量的，超过两个数量级。在4个具有6398特征的相同阵列上进行实验，每个阵列的平均变异系数（CV）为0.09，表明6398未处理的原始强度具有较高的再现性。在一个更精细的实验中，从无偏选择1125个酵母基因，发现在折叠变化水平上的平均CV为0.11。通过分析酵母对渗透冲击的转录反应，我们发现在我们的平台上获得的生物学数据与Affymetrix GeneChips的数据、定量实时PCR的数据以及从文献中获得的斑点cDNA阵列的数据非常一致，尽管有些不太清楚。
Here we describe a novel microarray platform that integrates all functions needed to perform any array-based experiment in a compact instrument on the researcher's laboratory benchtop. Oligonucle otide probes are synthesized in situ via a light- activated process within the channels of a three-dimensional microfluidic reaction carrier. Arrays can be designed and produced within hours according to the user's requirements. They are processed in a fully automatic workflow. We have characterized this new platform with regard to dynamic range, discrimination power, reproducibility and accuracy of biological results. The instrument detects sample RNAs present at a frequency of 1:100 000. Detection is quantitative over more than two orders of magnitude. Experiments on four identical arrays with 6398 features each revealed a mean coefficient of variation (CV) value of 0.09 for the 6398 unprocessed raw intensities indicating high reproducibility. In a more elaborate experiment targeting 1125 yeast genes from an unbiased selection, a mean CV of 0.11 on the fold change level was found. Analyzing the transcriptional response of yeast to osmotic shock, we found that biological data acquired on our platform are in good agreement with data from Affymetrix GeneChips, quantitative real-time PCR and-albeit somewhat less clearly-to data from spotted cDNA arrays obtained from the literature.