Publication Date:
2020-10-05
Description:
We report on the development of a microfluidic multiplexing technology for highly parallelized sample analysis via quantitative polymerase chain reaction (PCR) in an array of 96 nanoliter-scale microcavities made from silicon. This PCR array technology features fully automatable aliquoting microfluidics, a robust sample compartmentalization up to temperatures of 95 °C, and an application-specific prestorage of reagents within the 25 nl microcavities. The here presented hybrid silicon–polymer microfluidic chip allows both a rapid thermal cycling of the liquid compartments and a real-time fluorescence read-out for a tracking of the individual amplification reactions taking place inside the microcavities. We demonstrate that the technology provides very low reagent carryover of prestored reagents 〈 6 × 10−2 and a cross talk rate 〈 1 × 10−3 per PCR cycle, which facilitate a multi-targeted sample analysis via geometric multiplexing. Furthermore, we apply this PCR array technology to introduce a novel digital PCR-based DNA quantification method: by taking the assay-specific amplification characteristics like the limit of detection into account, the method allows for an absolute gene target quantification by means of a statistical analysis of the amplification results.
Print ISSN:
2096-1030
Electronic ISSN:
2055-7434
Topics:
Electrical Engineering, Measurement and Control Technology
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Energy, Environment Protection, Nuclear Power Engineering
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Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics
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