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研究了小牛胸腺脱氧核糖核酸(ctDNA)对硫堇(TH)荧光光谱的影响,发现DNA对TH的荧光有猝灭作用。对其猝灭机理进行了深入探讨,同时发现荧光猝灭度(ΔF)与DNA质量浓度(ρctDNA)之间存在线性关系。为消除Rayleigh散射干扰以建立良好的线性方程,选择TH荧光斯托克斯位移(4 nm)为Δλ进行同步荧光猝灭的研究,效果不佳。进一步考察了DNA对TH不同Δλ同步荧光的猝灭作用,分别选择λex/λem=627/638 nm和617/628 nm(Δλ=11 nm)作为参比溶液和样品溶液的工作波长,并建立了良好的线性方程,ctDNA的线性范围为0.015～1.260μg/mL,检出限为0.005μg/mL,回收率为96.5%～104.8%。该方法用于合成DNA样品的测定,结果满意。
The effect of calf thymus deoxyribonucleic acid (ctDNA) on the fluorescence spectrum of thionine (TH) was investigated. DNA was found to quench the fluorescence of TH. The mechanism of quenching was discussed in depth. At the same time, there was a linear relationship between fluorescence quenching (ΔF) and DNA mass concentration (ρctDNA). In order to eliminate the Rayleigh scattering interference and establish a good linear equation, the study of simultaneous fluorescence quenching using TH fluorescence Stokes shift (4 nm) as Δλ was not effective. Further investigation of the quenching effect of DNA on different Δλ synchrotron fluorescence of TH was done by using λex / λem = 627/638 nm and 617/628 nm (Δλ = 11 nm) as the working wavelengths of the reference solution and the sample solution respectively A good linear equation, ctDNA linear range of 0.015 ~ 1.260μg / mL, the detection limit of 0.005μg / mL, the recovery rate of 96.5% to 104.8%. The method was applied to the determination of DNA samples with satisfactory results.