Near-infrared fluorimetric determination of nucleic acids by shifting the ion-association equilibrium between heptamethylene cyanine and Alcian blue 8GX

摘要

A new method based on near-infrared (near-IR) fluorescence recovery, employing a two-reagent system which is composed of an anionic heptamethylene cyanine (HMC) and a polycationic phthalocyanine dye, Alcian blue 8GX, is presented for the determination of nucleic acids. With a maximum excitation wavelength at 766 nm and a maximum emission wavelength at 796 nm, the fluorescence recovery is linear with the concentration of nucleic acids added. Factors including the acidity of the medium, the reaction time, the optimal ratio of the two reagents, as well as the influence of foreign substance were all investigated. Meanwhile, the mechanism of fluorescence recovery was also studied. Under the optimal conditions, the linear ranges of the calibration curves were 10-250 nm ml~(-1) for calf thymus DNA (CT DNA) and 10-200 ng ml~(-1) for yeast RNA. The detection limits were 6.8 ng ml~(-1) for CT DNA and 6.3 ng ml~(-1) for yeast RNA, respectively. The method has been applied to the analysis of practical samples and the recovery results were satisfactory.