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Nucleic Acids Research, 2003, Vol. 31, No. 16 e89
© 2003 Oxford University Press

Gene expression analysis using single molecule detection

Kerstin Korn, Paola Gardellin1, Bohao Liao, Mario Amacker1, Åsa Bergström, Henrik Björkman, Agnès Camacho1, Sabine Dörhöfer, Klaus Dörre1, Johanna Enström1, Thomas Ericson, Tatiana Favez1, Michael Gösch, Adrian Honegger1, Sandra Jaccoud1, Markus Lapczyna, Erik Litborn, Per Thyberg, Holger Winter*,1 and Rudolf Rigler2

Gnothis AB, Electrum 212, SE-164 40 Kista, Sweden, 1 Gnothis SA, PSE-B EPFL CH-1015 Lausanne, Switzerland and 2 Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden

*To whom correspondence should be addressed. Tel: +41 21 693 8716; Fax: +41 21 693 8559; e-mail: Holger.Winter{at}gnothis.com

Recent developments of single molecule detection techniques and in particular the introduction of fluorescence correlation spectroscopy (FCS) led to a number of important applications in biological research. We present a unique approach for the gene expression analysis using dual-color cross-correlation. The expression assay is based on gene-specific hybridization of two dye-labeled DNA probes to a selected target gene. The counting of the dual-labeled molecules within the solution allows the quantification of the expressed gene copies in absolute numbers. As detection and analysis by FCS can be performed at the level of single molecules, there is no need for any type of amplification. We describe the gene expression assay and present data demonstrating the capacity of this novel technology. In order to prove the gene specificity, we performed experiments with gene-depleted total cDNA. The biological application was demonstrated by quantifying selected high, medium and low abundant genes in cDNA prepared from HL-60 cells.


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