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

Anke Pierik, M. Boamfa, M. Zelst, van, D. Clout, H.R. Stapert, J.F. Dijksman, D.J. Broer, R. Wimberger-Friedl

Research output: Contribution to journalArticleAcademicpeer-review

7 Citations (Scopus)
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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.
Original languageEnglish
Pages (from-to)18997-1902
Number of pages6
JournalLab on a Chip
Volume12
Issue number10
DOIs
Publication statusPublished - 2012

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