title: Microarray-based amplification and detection of RNA by nucleic acid sequence based amplification.
abstract: Nucleic acid sequence based amplification (NASBA) is a versatile in vitro nucleic acid amplification method. In this work, RNA amplification and labeling by NASBA and microarray analysis are combined in a one-step process. The NASBA reaction is performed in direct contact with capture probes. These probes are bound to surface-attached hydrogel spots generated at the chip surfaces by using a simple printing and UV irradiation process. Five gene expression and SNP parameters with known relevance in breast cancer diagnostics were chosen to demonstrate that multiplex NASBA-on-microarray analysis is possible. A minimum amount of 10 pg of total RNA was shown to be sufficient for the detection of the reference parameter RPS18, which demonstrates that the detection limit of the microarray-based NASBA assays theoretically allows single-cell assays to be performed.
authors.affiliation: Institute of Pathology, Freiburg University Medical Center, Breisacher Strasse 115a, 79106 Freiburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Mader
authors.forename: Ulrike
authors.initials: U
authors.lastname: Riehle
authors.forename: Thomas
authors.initials: T
authors.lastname: Brandstetter
authors.forename: Elmar
authors.initials: E
authors.lastname: Stickeler
authors.forename: Axel
authors.initials: A
authors.lastname: zur Hausen
authors.forename: Jürgen
authors.initials: J
authors.lastname: Rühe
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics; D005260:Female; D006367:HeLa Cells; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012313:RNA / Q000032:analysis* / Q000235:genetics; D021161:Self-Sustained Sequence Replication / Q000379:methods*
chemical_list: D012313:RNA