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Advances in DNA Detection on Paper Chips
KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
2013 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

DNA detection has an increasing importance in our everyday lives, with applications ranging from microbial diagnostics to forensic analysis. Currently, as the associated costs decrease, DNA diagnostic techniques are routinely used not only in research laboratories, but also in clinical and forensic practice.

The present thesis aims to unravel the potential of cellulose filter paper to be a viable candidate for DNA array support. There are two papers in this study. In Paper I, we studied the method of functionalizing the surface of filter paper and the possibility to detect DNA on acitve paper using fluorescence. In Paper II, we investigated visualization and throughput of DNA detection with magnetic beads on active filter papers, an assay which requires no instrumentation (scanner).

The findings in Paper I show that XG-NH2 and PDITC can functionalize the cellulose filter paper and that the activated filter papers can covalently bind oligonucleotides modified with amino groups to detect DNA. The detection limit of the assay is approximately 0.2 pmol. In Paper II, visualization of DNA detection on active paper is achieved without instrumentation, based on the natural color of magnetic beads. Furthermore, successful multiplex detection supports the potential to increase the throughput of DNA detection on active papers.

In summary, these studies show that active cellulose filter paper is a good DNA array support candidate as it provides a user-friendly and cost-efficient DNA detection assay. The methods described in Paper I and II are possible sources of development to a point-of-care device for on-site analysis of DNA contents in a sample.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2013. , 31 p.
Series
Trita-BIO-Report, ISSN 1654-2312 ; 2013:19
Keyword [en]
DNA detection, active filter papers, visualization, throughput, fluorescence, superparamagnetic beads
National Category
Other Industrial Biotechnology
Research subject
SRA - Molecular Bioscience
Identifiers
URN: urn:nbn:se:kth:diva-133614ISBN: 978-91-7501-922-2 (print)OAI: oai:DiVA.org:kth-133614DiVA: diva2:662591
Presentation
2013-11-29, Alfa 4, Kilsbergen, Science for Life Laboratory, Tomtebodavägen 23A, Stockholm, 14:00 (English)
Opponent
Supervisors
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 20131111

Available from: 2013-11-11 Created: 2013-11-07 Last updated: 2013-11-11Bibliographically approved
List of papers
1. Activated Paper Surfaces for the Rapid Hybridization of DNA through Capillary Transport
Open this publication in new window or tab >>Activated Paper Surfaces for the Rapid Hybridization of DNA through Capillary Transport
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2012 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 84, no 7, 3311-3317 p.Article in journal (Refereed) Published
Abstract [en]

The development of low-cost, accurate, and equipment-free diagnostic tests is crucial to many clinical, laboratory, and field applications, including forensics and medical diagnostics. Cellulose fiber-based paper is an inexpensive, biodegradable, and renewable resource, the use of which as a biomolecule detection matrix and support confers several advantages compared to traditional materials such as glass. In this context, a new, facile method for the preparation of surface functionalized papers bearing single-stranded probe DNA (ssDNA) for rapid target hybridization via capillary transport is presented. Optimized reaction conditions were developed that allowed the direct, one-step activation of standard laboratory filters by the inexpensive and readily available bifunctional linking reagent, 1,4-phenylenediisothiocyanate. Such papers were thus amenable to subsequent coupling of amine-labeled ssDNA under standard conditions widely used for glass-based supports. The intrinsic wicking ability of the paper matrix facilitated rapid sample elution through arrays of probe DNA, leading to significant, detectable hybridization in the time required for the sample liquid to transit the vertical length of the strip (less than 2 min). The broad applicability of these paper test strips as rapid and specific diagnostics in "real-life" situations was exemplified by the discrimination of amplicons generated from canine and human mitochondrial and genomic DNA in mock forensic samples.

Keyword
Bioactive Paper, Visual Dna, Cellulose, Xyloglucan, Biosensors, Aptamers, Assay
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-95103 (URN)10.1021/ac300025v (DOI)000302829800039 ()22369042 (PubMedID)2-s2.0-84859392980 (Scopus ID)
Funder
Swedish Research CouncilKnut and Alice Wallenberg FoundationScience for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 20150629

Available from: 2012-05-21 Created: 2012-05-14 Last updated: 2017-12-07Bibliographically approved
2. Visual detection of DNA on paper chips
Open this publication in new window or tab >>Visual detection of DNA on paper chips
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2014 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 86, no 3, 1575-1582 p.Article in journal (Refereed) Published
Abstract [en]

On-site DNA analysis for diagnostic or forensic purposes is much anticipated in the future of molecular testing. Yet the challenges to achieve this goal remain large with rapid and inexpensive detection and visualization being key factors for any portable analysis system. We have developed a filter paper-based nucleic acid assay, which is able to identify and distinguish dog and human genomic and mitochondrial samples in a forensic setting. The filter paper material allows for transport by capillary force of the sample DNA through the detection surface, allowing the targets to hybridize specifically to their complementary capture sequences. Coupling micrometer-sized beads to DNA allows the results to be visualized by the naked eye, enabling instant, cost-efficient, and on-site detection, while eliminating the need for advanced expensive instrumentation.

Keyword
Analysis system, Capillary force, Cost-efficient, DNA analysis, Filter paper material, Nucleic acid assays, On-site detection, Visual detection
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-133777 (URN)10.1021/ac403196b (DOI)000331014800038 ()2-s2.0-84893634724 (Scopus ID)
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscienceSwedish Research CouncilFormas
Note

QC 20140313. Updated from manuscript to article in journal.

Available from: 2013-11-11 Created: 2013-11-11 Last updated: 2017-12-06Bibliographically approved

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