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Multiplex PCB-based electrochemical detection of cancer biomarkers using MLPA-barcode approach
KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
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2016 (English)In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 82, 224-232 p.Article in journal (Refereed) PublishedText
Abstract [en]

Asymmetric multiplex ligation-dependent probe amplification (MLPA) was developed for the amplification of seven breast cancer related mRNA markers and the MLPA products were electrochemically detected via hybridization. Seven breast cancer genetic markers were amplified by means of the MLPA reaction, which allows for multiplex amplification of multiple targets with a single primer pair. Novel synthetic MLPA probes were designed to include a unique barcode sequence in each amplified gene. Capture probes complementary to each of the barcode sequences were immobilized on each electrode of a low-cost electrode microarray manufactured on standard printed circuit board (PCB) substrates. The functionalised electrodes were exposed to the single-stranded MLPA products and following hybridization, a horseradish peroxidase (HRP)-labelled DNA secondary probe complementary to the amplified strand completed the genocomplex, which was electrochemically detected following substrate addition. The electrode arrays fabricated using PCB technology exhibited an excellent electrochemical performance, equivalent to planar photolithographically-fabricated gold electrodes, but at a vastly reduced cost (>50 times lower per array). The optimised system was demonstrated to be highly specific with negligible cross-reactivity allowing the simultaneous detection of the seven mRNA markers, with limits of detections as low as 25 pM. This approach provides a novel strategy for the genetic profiling of tumour cells via integrated "amplification-to-detection".

Place, publisher, year, edition, pages
Elsevier, 2016. Vol. 82, 224-232 p.
Keyword [en]
Barcodes, Breast cancer markers, Electrochemical detection, Multiplex ligation-dependent probe amplification system (MLPA), PCB genosensor arrays, Amplification, Bar codes, Biomarkers, Chemical detection, Diseases, Electrodes, Gain control, Multiplexing, Nucleic acids, Printed circuit boards, Printed circuits, Probes, Substrates, Breast Cancer, Electrochemical performance, Genosensors, Horseradish peroxidase, Printed circuit boards (PCB), Probe amplification, Simultaneous detection, Electrochemical electrodes
National Category
Cancer and Oncology Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:kth:diva-186895DOI: 10.1016/j.bios.2016.04.018ScopusID: 2-s2.0-84962802416OAI: oai:DiVA.org:kth-186895DiVA: diva2:930558
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QC 20160524

Available from: 2016-05-24 Created: 2016-05-16 Last updated: 2016-05-24Bibliographically approved

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Solnestam, Beata WerneKvastad, LindaAkan, PelinLundeberg, Joakim
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Gene TechnologyScience for Life Laboratory, SciLifeLab
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Cancer and OncologyMedical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)

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