title: Spotting and validation of a genome wide oligonucleotide chip with duplicate measurement of each gene.
abstract: The quality of DNA microarray based gene expression data relies on the reproducibility of several steps in a microarray experiment. We have developed a spotted genome wide microarray chip with oligonucleotides printed in duplicate in order to minimise undesirable biases, thereby optimising detection of true differential expression. The validation study design consisted of an assessment of the microarray chip performance using the MessageAmp and FairPlay labelling kits. Intraclass correlation coefficient (ICC) was used to demonstrate that MessageAmp was significantly more reproducible than FairPlay. Further examinations with MessageAmp revealed the applicability of the system. The linear range of the chips was three orders of magnitude, the precision was high, as 95% of measurements deviated less than 1.24-fold from the expected value, and the coefficient of variation for relative expression was 13.6%. Relative quantitation was more reproducible than absolute quantitation and substantial reduction of variance was attained with duplicate spotting. An analysis of variance (ANOVA) demonstrated no significant day-to-day variation.
authors.affiliation: Department of Biochemistry, Pharmacology, and Genetics, Odense University Hospital and Human Microarray Centre, University of Southern Denmark, Odense, Denmark. mads.thomassen@ouh.fyns-amt.dk
authors.forename: Mads
authors.initials: M
authors.lastname: Thomassen
authors.forename: Vibe
authors.initials: V
authors.lastname: Skov
authors.forename: Freyja
authors.initials: F
authors.lastname: Eiriksdottir
authors.forename: Qihua
authors.initials: Q
authors.lastname: Tan
authors.forename: Kirsten
authors.initials: K
authors.lastname: Jochumsen
authors.forename: Niels
authors.initials: N
authors.lastname: Fritzner
authors.forename: Klaus
authors.initials: K
authors.lastname: Brusgaard
authors.forename: Jesper
authors.initials: J
authors.lastname: Dahlgaard
authors.forename: Torben A
authors.initials: TA
authors.lastname: Kruse
mesh_terms: D020869:Gene Expression Profiling / Q000379:methods*; D016678:Genome / Q000235:genetics*; D015894:Genome, Human / Q000235:genetics; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D020161:Physical Chromosome Mapping / Q000379:methods*