Validation of a novel, fully integrated and flexible microarray benchtop facility for gene expression profiling.
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.
Baum
Michael
M
febit ag, Käfertaler Strasse 190, 68167 Mannheim, Germany. michael.baum@febit.de
Bielau
Simone
S
Rittner
Nicole
N
Schmid
Kathrin
K
Eggelbusch
Kathrin
K
Dahms
Michael
M
Schlauersbach
Andrea
A
Tahedl
Harald
H
Beier
Markus
M
Güimil
Ramon
R
Scheffler
Matthias
M
Hermann
Carsten
C
Funk
Jörg-Michael
JM
Wixmerten
Anke
A
Rebscher
Hans
H
Hönig
Matthias
M
Andreae
Claas
C
Büchner
Daniel
D
Moschel
Erich
E
Glathe
Andreas
A
Jäger
Evelyn
E
Thom
Marc
M
Greil
Andreas
A
Bestvater
Felix
F
Obermeier
Frank
F
Burgmaier
Josef
J
Thome
Klaus
K
Weichert
Sigrid
S
Hein
Silke
S
Binnewies
Tim
T
Foitzik
Volker
V
Müller
Manfred
M
Stähler
Cord Friedrich
CF
Stähler
Peer Friedrich
PF
D001331:Automation / Q000295:instrumentation; D020869:Gene Expression Profiling / Q000295:instrumentation*; D005800:Genes, Fungal / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation*; D012331:RNA, Fungal / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000235:genetics; D012680:Sensitivity and Specificity
D012331:RNA, Fungal; D012333:RNA, Messenger