Quantitative assessment of a novel flow-through porous microarray for the rapid analysis of gene expression profiles.
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.
Wu
Ying
Y
PamGene International BV, Nieuwstraat 30, PO Box 1345, 5200 BJ 's-Hertogenbosch, The Netherlands. ywu@pamgene.com
de Kievit
Peggy
P
Vahlkamp
Lars
L
Pijnenburg
Dirk
D
Smit
Maarten
M
Dankers
Martijn
M
Melchers
Diana
D
Stax
Martijn
M
Boender
Piet J
PJ
Ingham
Colin
C
Bastiaensen
Niek
N
de Wijn
Rik
R
van Alewijk
Dirk
D
van Damme
Henk
H
Raap
Anton K
AK
Chan
Alan B
AB
van Beuningen
Rinie
R
D000537:Aluminum Oxide / Q000737:chemistry; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015203:Reproducibility of Results; D013997:Time Factors
D000537:Aluminum Oxide