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Use of Surface Plasmon-Coupled Emission to Measure DNA Hybridization

Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology

Ignacy Gryczynski

Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology

Zygmunt Gryczynski

Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology

Joseph R. Lakowicz

Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology

The authors describe a new approach to measuring DNA hybridization based on surface plasmon-coupled emission (SPCE). SPCE is the resonance coupling of excited fluorophores with electron motions in thin metal films, resulting in efficient transfer of energy through the film and radiation into the glass substrate. The authors evaluated the use of SPCE for detection of DNA hybridization. An unlabeled capture biotinylated oligonucleotide was attached near the surface of a thin (50 nm) silver film using streptavidin. The authors then measured the emission intensity of single-stranded Cy5-labeled DNA upon binding to a complementary oligomer attached to a silver film. Hybridization could be detected by an increase in SPCE, which appeared as light radiated into the substrate at a sharply defined angle near 73 degrees from the normal. The largest signals were observed when the excitation angle of incidence equaled the surface plasmon wavelength, but directional emission was also observed without excitation by the surface plasmon evanescent field. The increased intensity is due to proximity to the metal surface, so that hybridization can be detected without a change in the quantum yield of the fluorophore. These results indicate that SPCE can provide highly sensitive real-time measurement of DNA hybridization.

Key Words: PDNA microarrays • DNA hybridization • fluorescence • surface plasmons • surface plasmon resonance • directional emission

Journal of Biomolecular Screening, Vol. 9, No. 3, 208-215 (2004)
DOI: 10.1177/1087057103262363


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