Portal de Periódicos da CAPES

Validation of a novel, fully integrated and flexible microarray benchtop facility for gene expression profiling

2003; Oxford University Press; Volume: 31; Issue: 23 Linguagem: Inglês

10.1093/nar/gng151

1362-4962

Michaël Baum, Simone Bielau, Nicole Rittner, Kathrin Schmid, Kathrin Eggelbusch, Michael Dahms, Andrea Schlauersbach, Harald Tahedl, Markus Beier, Ramon Güimil, Matthias Scheffler, Carsten Hermann, J.A. Funk, Anke Wixmerten, Hans Rebscher, Matthias Hönig, Claas Andreae, Daniel Büchner, Erich Moschel, Andreas Glathe, Evelyn Jäger, M. Thom, Andreas Greil, Felix Bestvater, Frank Obermeier, Josef Burgmaier, Klaus Thome, Sigrid Weichert, Silke Hein, Tim T. Binnewies, Volker Foitzik, Manfred Müller, Cord Stähler, Peer Friedrich Stähler,

Molecular Biology Techniques and Applications

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.