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Issue 10, 2012
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Real time quantitative amplification detection on a microarray: towards high multiplex quantitative PCR

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Abstract

Quantitative real-time polymerase chain reaction (qrtPCR) is widely used as a research and diagnostic tool. Notwithstanding its many powerful features, the method is limited in the degree of multiplexing to about 6 due to spectral overlap of the available fluorophores. A new method is presented that allows quantitative amplification detection at higher multiplexing by the integration of amplification in solution and monitoring via hybridization to a microarray in real-time. This method does not require any manipulation of the PCR product and runs in a single closed chamber. Employing labeled primers, one of the main challenges is to measure surface signals against a high fluorescence background from solution. A compact, confocal scanner is employed, based on miniaturized optics from DVD technology and combined with a flat thermocycler for simultaneous scanning and heating. The feasibility of this method is demonstrated in singleplex with an analytical sensitivity comparable to routine qrtPCR.

Graphical abstract: Real time quantitative amplification detection on a microarray: towards high multiplex quantitative PCR

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Publication details

The article was received on 10 Aug 2011, accepted on 24 Feb 2012 and first published on 03 Apr 2012


Article type: Technical Note
DOI: 10.1039/C2LC20740K
Citation: Lab Chip, 2012,12, 1897-1902
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    Real time quantitative amplification detection on a microarray: towards high multiplex quantitative PCR

    A. Pierik, M. Boamfa, M. van Zelst, D. Clout, H. Stapert, F. Dijksman, D. Broer and R. Wimberger-Friedl, Lab Chip, 2012, 12, 1897
    DOI: 10.1039/C2LC20740K

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