title: RNA expression profiling at the single molecule level.
abstract: We developed a microarray platform for PCR amplification-independent expression profiling of minute samples. A novel scanning system combined with specialized biochips enables detection down to individual fluorescent oligonucleotide molecules specifically hybridized to their complementary sequence over the entire biochip surface of cm2 size. A detection limit of 1.3 fM target oligonucleotide concentration--corresponding to only 39,000 molecules in the sample solution--and a dynamic range of 4.7 orders of magnitude have been achieved. The applicability of the system to PCR amplification-independent gene-expression profiling of minute samples was demonstrated by complex hybridization of cDNA derived from the equivalent of only 10(4) cells, which matches results obtained in ensemble studies on large samples. By counting each hybridized molecule on the microarray, the method is insusceptible to gene-specific variations of the labeling, thereby representing a principle advance to conventional ensemble-based microarray analysis.
authors.affiliation: Biophysics Institute, Johannes Kepler University Linz, A-4040 Linz, Austria.
authors.forename: Jan
authors.initials: J
authors.lastname: Hesse
authors.forename: Jaroslaw
authors.initials: J
authors.lastname: Jacak
authors.forename: Maria
authors.initials: M
authors.lastname: Kasper
authors.forename: Gerhard
authors.initials: G
authors.lastname: Regl
authors.forename: Thomas
authors.initials: T
authors.lastname: Eichberger
authors.forename: Martina
authors.initials: M
authors.lastname: Winklmayr
authors.forename: Fritz
authors.initials: F
authors.lastname: Aberger
authors.forename: Max
authors.initials: M
authors.lastname: Sonnleitner
authors.forename: Robert
authors.initials: R
authors.lastname: Schlapak
authors.forename: Stefan
authors.initials: S
authors.lastname: Howorka
authors.forename: Leila
authors.initials: L
authors.lastname: Muresan
authors.forename: Anna-Maria
authors.initials: AM
authors.lastname: Frischauf
authors.forename: Gerhard J
authors.initials: GJ
authors.lastname: Schütz
mesh_terms: D002460:Cell Line; D015342:DNA Probes; D018076:DNA, Complementary; D020869:Gene Expression Profiling; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction
chemical_list: D015342:DNA Probes; D018076:DNA, Complementary