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Pinched flow fractionation devices for detection of single nucleotide polymorphisms

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Authors:
  • Larsen, Asger Vig ;
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    Department of Micro- and Nanotechnology, Technical University of Denmark
  • Poulsen, Lena ;
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    Department of Micro- and Nanotechnology, Technical University of Denmark
  • Birgens, Henrik ;
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    unknown
  • Dufva, Hans Martin ;
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    Orcid logo0000-0001-5449-0189
    Department of Micro- and Nanotechnology, Technical University of Denmark
  • Kristensen, Anders
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    Department of Micro- and Nanotechnology, Technical University of Denmark
DOI:
10.1039/b802268b
Abstract:
We demonstrate a new and flexible micro fluidic based method for genotyping single nucleotide polymorphisms ( SNPs). The method relies on size separation of selectively hybridized polystyrene microspheres in a micro fluidic pinched flow fractionation (PFF) device. The micro fluidic PFF devices with 13 mu m deep channels were fabricated by thermal nanoimprint lithography ( NIL) in a thin film of cyclic-olefin copolymer (mr-I T85) on a silicon wafer substrate, and the channels were sealed by thermal polymer bonding. Streptavidin coated polystyrene microspheres with a mean diameter of 3.09 mu m and 5.6 mu m were functionalized with biotin-labeled oligonucleotides for the detection of a mutant (Mt) or wild-type (Wt) DNA sequence in the HBB gene, respectively. Hybridization to functionalized beads was performed with fluorescent targets comprising synthetic DNA oligonucleotides or amplified RNA, synthesized using human DNA samples from individuals with point mutations in the HBB gene. Following a stringent wash, the beads were separated in a PFF device and the fluorescent signal from the beads was analyzed. Patients being wildtypes, heterozygotes or mutated respectively for the investigated mutation could reliably be diagnosed in the PFF device. This indicates that the PFF technique can be used for accurate and fast genotyping of SNPs.
Type:
Journal article
Language:
English
Published in:
Lab on a Chip, 2008, Vol 8, Issue 5, p. 818-821
Keywords:
Main Research Area:
Science/technology
Publication Status:
Published
Review type:
Peer Review
Submission year:
2008
Scientific Level:
Scientific
ID:
22559652

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