Nucleic Acids Analysis with Nano-Ag-Tb(III) by a Resonance Light Scattering Technique

Abstract

The quantitative analysis of nucleic acids is very important because it can be used as a reference for measurements of other components in biological fluids and genetic diagnosis. Up to now, various analytical methods have been developed for the determination of nucleic acids, such as fluorometry, 1,2 chemiluminescence, 3 and electrochemistry. 4 Recently, fluorescent probes including organic dyes, such as yellow orange dyes, 5,6 metal ions, such as Tb(III), 7 and a metal complex 8 are frequently employed to investigate nucleic acids.Rayleigh scattering is a light-scattering phenomenon where electromagnetic radiation and a material interact with each other and generate elastic collisions.Based on macroscopic fluctuation theory, the intensity of scattered Rayleigh light is in proportion to λ0 -4 , 9 since the light scattering of molecular particles is 20-fold smaller than the wavelength of the incident beam. If the wavelength of the incident beam is close to that of the absorption band of the molecular particles, Rayleigh scattering will deviate from the ordinary law, and the intensity of some wavelengths will rapidly increase. This phenomenon is called resonance light scattering (RLS investigated the enhancing and quenching functions of nano-Ag on the luminescent properties of an europium complex, in which the effects of particles upon aggregation were considered.In this paper, the RLS of nano-Ag-Tb(III)-nucleic acids system is reported for the first time. The results show that with a common spectrofluorometer and inexpensive reagents, nucleic acids can be detected by this method with satisfactory results. Experimental ApparatusA transmission electron microscopy (TEM) image of the nano-Ag was acquired using a Hitachi H-600 (Japan) transmission electron microscope. The RLS spectrum and the intensity of RLS were measured with a Hitachi 850 spectrofluorometer (Japan).All absorption spectra were recorded with a Cintra 10e UV-Vis spectrophotometer (GBC). A 420A plus pH meter (Orion Research Inc.) was used to measure the pH of the solutions. ReagentsStock solutions of nucleic acids (100 µg ml -1 ) were prepared by dissolving purchased fish sperm DNA (fsDNA), calf thymus DNA (ctDNA) or yeast RNA (yRNA) (Sigma) in water. These stocks were stored at 0 -4˚C. The concentrations of nucleic acids, where necessary, were calculated according to the absorbance at 260 nm by using the molar extinction coefficients, εDNA (6600 l mol -1 cm -1 ) and εRNA (7800 l mol -1 cm -1 ), after establishing that the absorbance ratio, A260/A280, was 1.8 for DNA and 2.0 for RNA, respectively. A standard stock solution of the Tb(III) ion (0.01 mol l -1 ) was prepared by dissolving 934.5 mg of Tb4O7 in 15 ml of HCl (12 mol l -1 ) at 100˚C, and evaporating the solution to be almost dry; it was then diluted to 500 ml with doubly distilled water. The nano-Ag-terbium(III)-nucleic acids system was observed by a resonance light scattering (RLS) technique for the first time, and the quantitative analysis of nucleic acids at nanogram levels was establishe...

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