title: A common reference for cDNA microarray hybridizations.
abstract: Comparisons of expression levels across different cDNA microarray experiments are easier when a common reference is co-hybridized to every microarray. Often this reference consists of one experimental control sample, a pool of cell lines or a mix of all samples to be analyzed. We have developed an alternative common reference consisting of a mix of the products that are spotted on the array. Pooling part of the cDNA PCR products before they are printed and their subsequent amplification towards either sense or antisense cRNA provides an excellent common reference. Our results show that this reference yields a reproducible hybridization signal in 99.5% of the cDNA probes spotted on the array. Accordingly, a ratio can be calculated for every spot, and expression levels across different hybridizations can be compared. In dye-swap experiments this reference shows no significant ratio differences, with 95% of the spots within an interval of +/-0.2-fold change. The described method can be used in hybridizations with both amplified and non-amplified targets, is time saving and provides a constant batch of common reference that lasts for thousands of hybridizations.
authors.affiliation: Center for Human and Clinical Genetics, Leiden University Medical Center, Wassenaarseweg 72, 2333AL Leiden, Nederland.
authors.forename: Ellen
authors.initials: E
authors.lastname: Sterrenburg
authors.forename: Rolf
authors.initials: R
authors.lastname: Turk
authors.forename: Judith M
authors.initials: JM
authors.lastname: Boer
authors.forename: Gertjan B
authors.initials: GB
authors.lastname: van Ommen
authors.forename: Johan T
authors.initials: JT
authors.lastname: den Dunnen
mesh_terms: D002478:Cells, Cultured; D005347:Fibroblasts; D015723:Gene Library; D006801:Humans; D009132:Muscles; D009693:Nucleic Acid Hybridization / Q000379:methods; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods* / Q000592:standards*; D016133:Polymerase Chain Reaction; D011786:Quality Control; D012015:Reference Standards; D015203:Reproducibility of Results; D013997:Time Factors