Two single-stranded DNAs consisting of complementary base pairs except for one mismatching base pair (MM1) can form double-stranded DNA by molecular recognition. This type of duplex is not as stable as that formed by MM0. In order to add to a better understanding of the physical mechanism of the hybridization and dissociation processes at sensor (chip) surfaces, we studied the kinetics of the MM1 hybridization by surface plasmon fluorescence spectroscopy. Target DNA strands labelled with a fluorescent molecule Cy5 at the 5' end and hybridizing with the surface-attached probe DNA can be excited by the strong optical field of a surface plasmon resonance mode. The emitted fluorescence can be detected with high sensitivity. The affinity of a duplex was found to depend on the chemical nature, i.e. G-G, G-T etc., and on the position of the mismatching base pair along the 15mer duplex.

On the substrate carrying a sub-wavelength grating covered with a thin metal layer, a fluorescent dye-labeled cell was observed by fluorescence microscope. The fluorescence intensity was more than 20 times greater than that on an optically flat glass substrate. Such a great fluorescence enhancement from labeled cells bound to the grating substrate was due to the excitation by grating coupled surface plasmon resonance. The application of a grating substrate to two-dimensional detection and fluorescence microscopy appears to offer a promising method of taking highly sensitive fluorescence images.