Studies On The Determination Of DNA By Resonance Light Scattering Spectrometry

In this thesis, we summarize the development, discovery, and basically principleof resonance light scattering (RLS). The double-stranded DNA (dsDNA) and30-mersingle-stranded DNA (ssDNA) were determined by RLS.When aggregate is irradiated with a light beam with the wavelength close totheir absorption bands of the aggregate, the enhanced RLS signals can be easilymeasured by scanning the excitation and the emission monochromators of a commonspectrofluorometer simultaneously. RLS signals are related to the structure, shape,size, charge distribution and refractive index (RI) of the surrounding medium of themolecule. RLS spectrometry has widely applied to the determination of metal ions,medicines and proteins, especially DNA.The RLS spectrometry of determining DNA is based on the shifts of RLSintensity owing to the interaction between DNA and RLS probe. There are manymolecule probes for the determination of DNA, such as the metal complexes,nanoparticles (NPs), and cationic surfactants.Gold nanostructures, especially nonspherical gold nanostructures, have beenapplied to numerous fields and received the great attention because their opticalproperties depend on the shape and size of the nanostructures. Nonspherical goldnanostructures usually exhibit multiple surface plasmon-resonance bands that aretunable over a broad spectral range from the visible to NIR region, which result from dipolar and multipolar electron oscillations along different directions. For example,gold bipyramids and gold nanorods have two distinct surface plasmon-resonancebands. Gold bipyramids have a pentagonal base and two sharp apexes. Goldbipyramids possess transverse and longitudinal surface plasmon resonance and thelatter can be tuned by adjusting the ratio of the height to the base-edge length.Because the apexes of gold bipyramids are sharper than the ends of gold nanorods, thelocal electric-field enhancements associated with bipyramids should be larger thanthat associated with nanorods, according to the lightning-rod effect.The fsDNA was determined by RLS spectrometry in the second chapter. Thegold bipyramids were synthesized by a seed-mediated method and used as the probes.Cetyltrimethylammonium bromide (CTAB) was used as stabilizing agent. Thenegative charged fsDNA is bound to cationic surfactant CTAB on the surface of thegold bipyramids to form complexes by electrostatic adsorption, and significantchanges occur in the plasmon spectra in response to RI changes in the vicinity of thecomplexes, which results in a significant enhancement of the RLS intensity.According to the RLS theory, the RLS signal is enhanced when the size of scatteringparticle is enlarged. The size of complex of gold bipyramid-fsDNA is much largerthan that of gold bipyramids, so the RLS intensity increases drastically. Under theoptimal conditions, the intensity of RLS is directly proportional to the concentrationof DNA in the range from0.1to2.0μg/mL.Based on the aggregation of gold bipyramids, a novel RLS biosensor fordetermining sequence-specific DNA was developed in the third chapter. When theprobe DNA was added into the gold bipyramid suspension, probe DNA was adsorbedonto the surface of gold bipyramids, and the gold bipyramid-ssDNA complex wasformed. After the target DNA which is complementary to probe DNA is added into thegold bipyramid-ssDNA complex suspension, DNA hybridization is first completed asa consequence of the formation of dsDNA, because the charge density of dsDNA ismuch larger than that of ssDNA. Therefore, the dsDNA molecules might act as “glue”to stick the gold bipyramids together by the CTAB bilayer one by one through strongelectrostatic interaction to induce aggregation of gold bipyramids. The present method was successfully applied to the determination of a30-mer single-strandedoligonucleotide in the concentration range of0.1–10nmol/L.