AVS 47th International Symposium
    Biomaterial Interfaces Friday Sessions
       Session BI-FrM

Paper BI-FrM4
PNA-DNA and DNA-DNA Hybridization Detection via Lipid-Biotin-Streptavidin Immobilization on a SiO2 Coated Quartz Crystal Microbalance Sensor

Friday, October 6, 2000, 9:20 am, Room 202

Session: Biomolecular Recognition at Surfaces
Presenter: F. Höök, Chalmers University of Technology, Sweden
Authors: F. Höök, Chalmers University of Technology, Sweden
A. Ray, Chalmers University of Technology, Sweden
B. Norden, Chalmers University of Technology, Sweden
B. Kasemo, Chalmers University of Technology, Sweden
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Surface-based bioanalytical sensors for oligonucleotide hybridization are very attractive for genetic diagnostics, gene therapeutics and the emerging solid phase / real time PCR. Little is however known about how various immobilization strategies affect the conformation and hence function of oligonucleotide strands. We have investigated the possibility to use the dissipative quartz crystal microbalance (QCM-D) technique and controlled surface-immobilization of single stranded synthetic peptide nucleic acid (PNA) as well as DNA, as selective probe(s) for fully complementary and various single mismatch DNA. In order to minimize unspecific binding of DNA, streptavidin was immobilized as a protein 2-D crystal on a biotinylated phospholipid bilayer supported on a SiO@sub 2@ surface in the fluid liquid crystalline phase. This was followed by attachment of a mixed-sequence 15-mer biotin-PNA or a 15-mer biotin-DNA with identical base pairs. The exposure of the streptavidin-immobilized biotin-PNA and biotin-DNA to fully complementary and various mismatch DNA was investigated at 24 °C. Only the fully complementary and singly mismatched DNA oligomers hybridized with the immobilized PNA and DNA, and was possible to discriminate via significant difference in the binding and dissociation kinetics, demonstrating a very high selectively. Most interestingly, however, is that the ratio between hybridization-induced energy dissipation (c.f. rigidity) and the frequency shifts (c.f. mass uptake), allowed us to discriminate different structures of immobilized PNA-DNA and DNA-DNA duplexes. Possible influence on the hybridization kinetics and the structure of the formed duplexes from primarily lateral interaction is discussed.