Towards a “Sample-In, Answer-Out” Point-of-Care Platform for Nucleic Acid Extraction and Amplification: Using an HPV E6/E7 mRNA Model System

Author:

Gulliksen Anja12,Keegan Helen3,Martin Cara3,O'Leary John3,Solli Lars A.1,Falang Inger Marie1,Grønn Petter1,Karlgård Aina1,Mielnik Michal M.4,Johansen Ib-Rune4,Tofteberg Terje R.24,Baier Tobias56,Gransee Rainer5,Drese Klaus5,Hansen-Hagge Thomas5,Riegger Lutz78,Koltay Peter78,Zengerle Roland7,Karlsen Frank19,Ausen Dag4,Furuberg Liv24

Affiliation:

1. NorChip AS, Industriveien 8, 3490 Klokkarstua, Norway

2. University of Oslo, 0316 Oslo, Norway

3. Department of Histopathology, Trinity College Dublin and Molecular Pathology Research Laboratory, Coombe Women and Infants University Hospital, Dolphins Barn, Dublin 8, Ireland

4. SINTEF ICT, MiNaLab Facility, Gaustadalléen 23C, 0373 Oslo, Norway

5. Institut für Mikrotechnik Mainz, Carl-Zeiss Straße 18-20, 55129 Mainz, Germany

6. Center of Smart Interfaces, TU Darmstadt, Petersenstraße 32, 64287 Darmstadt, Germany

7. Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Georges-Koehler-Allee 106, 79110 Freiburg, Germany

8. BioFluidix GmbH, Georges-Koehler-Allee 106, 79110 Freiburg, Germany

9. Department of Micro and Nano Systems Technology, Faculty of Technology and Maritime Sciences, Vestfold University College, Raveien 197, 3184 Borre, Norway

Abstract

The paper presents the development of a “proof-of-principle” hands-free and self-contained diagnostic platform for detection of human papillomavirus (HPV) E6/E7 mRNA in clinical specimens. The automated platform performs chip-based sample preconcentration, nucleic acid extraction, amplification, and real-time fluorescent detection with minimal user interfacing. It consists of two modular prototypes, one for sample preparation and one for amplification and detection; however, a common interface is available to facilitate later integration into one single module. Nucleic acid extracts (n=28) from cervical cytology specimens extracted on the sample preparation chip were tested using the PreTect HPV-Proofer and achieved an overall detection rate for HPV across all dilutions of 50%–85.7%. A subset of 6 clinical samples extracted on the sample preparation chip module was chosen for complete validation on the NASBA chip module. For 4 of the samples, a 100% amplification for HPV 16 or 33 was obtained at the 1 : 10 dilution for microfluidic channels that filled correctly. The modules of a “sample-in, answer-out” diagnostic platform have been demonstrated from clinical sample input through sample preparation, amplification and final detection.

Funder

European Commission

Publisher

Hindawi Limited

Subject

Oncology

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