Digital Droplet PCR on Disk

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Abstract: Existing systems for digital droplet PCR (ddPCR) either suffer from low integration or are difficult to introduce to mass fabrication. Here we present an integrated system that is compatible to mass fabrication and combines emulsification, PCR, and fluorescence readout in a single chamber within a disposable cartridge (disk). Droplets are generated by injecting the sample into fluorinated oil via centrifugal step emulsification. The resulting emulsion is aligned in the PCR and readout zone by capillary action. During thermocycling, gas bubbles generated by degassing are removed by capillary driven transport through tapered regions in the PCR chamber. Thereby, the positioning of the emulsion within the readout zone of the PCR chamber is ensured at any time and no bubbles are present during readout. Manual handling of the disk solely requires pipetting of oil and PCR mix into the inlet structures, placing the disk into the thermocycler and subsequently into a microarray scanner. The functionality of the ddPCR process chain is demonstrated by quantitative detection of the cystic fibrosis causing mutation p.Phe508del, which is of interest for non-invasive prenatal testing (NIPT). The mutation was detected in a concentration range spanning four orders of magnitude. We envision that this work will lay the base for the development of highly integrated sample-to-digital-answer PCR systems that can be employed in routine clinical diagnosis

Location
Deutsche Nationalbibliothek Frankfurt am Main
Extent
Online-Ressource
Language
Englisch
Notes
Lab on a chip. 16(2016), 208-216, DOI 10.1039/C5LC01068C, issn: 1473-0189
IN COPYRIGHT http://rightsstatements.org/page/InC/1.0 rs

Classification
Physik

DOI
10.1039/C5LC01068C
URN
urn:nbn:de:bsz:25-freidok-128087
Rights
Der Zugriff auf das Objekt ist unbeschränkt möglich.
Last update
25.03.2025, 1:41 PM CET

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Time of origin

  • 2015

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