| Literature DB >> 15333674 |
Ying Wu1, Peggy de Kievit, Lars Vahlkamp, Dirk Pijnenburg, Maarten Smit, Martijn Dankers, Diana Melchers, Martijn Stax, Piet J Boender, Colin Ingham, Niek Bastiaensen, Rik de Wijn, Dirk van Alewijk, Henk van Damme, Anton K Raap, Alan B Chan, Rinie van Beuningen.
Abstract
A novel microarray system that utilizes a porous aluminum-oxide substrate and flow-through incubation has been developed for rapid molecular biological testing. To assess its utility in gene expression analysis, we determined hybridization kinetics, variability, sensitivity and dynamic range of the system using amplified RNA. To show the feasibility with complex biological RNA, we subjected Jurkat cells to heat-shock treatment and analyzed the transcriptional regulation of 23 genes. We found that trends (regulation or no change) acquired on this platform are in good agreement with data obtained from real-time quantitative PCR and Affymetrix GeneChips. Additionally, the system demonstrates a linear dynamic range of 3 orders of magnitude and at least 10-fold decreased hybridization time compared to conventional microarrays. The minimum amount of transcript that could be detected in 20 microl volume is 2-5 amol, which enables the detection of 1 in 300,000 copies of a transcript in 1 microg of amplified RNA. Hybridization and subsequent analysis are completed within 2 h. Replicate hybridizations on 24 identical arrays with two complex biological samples revealed a mean coefficient of variation of 11.6%. This study shows the potential of flow-through porous microarrays for the rapid analysis of gene expression profiles in clinical applications.Entities:
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Year: 2004 PMID： 15333674 PMCID： PMC516077 DOI： 10.1093/nar/gnh118
Source DB: PubMed Journal: Nucleic Acids Res ISSN： 0305-1048 Impact factor: 16.971