荧光标记DNA高分辨电感耦合等离子体质谱定量分析  

Quantitative Analysis of Fluorescent Dye-labeled DNA by High Resolution Inductively Coupled Plasma Mass Spectrometry

作　　者：高运华[1] 李海峰[1] 李建新 黎朋[1] 叶健[3] 赵兴春[3] 

机构地区：[1]中国计量科学研究院,北京100013 [2]菏泽市产品质量监督检验所,菏泽274000 [3]公安部物证鉴定中心,北京100038

出　　处：《高等学校化学学报》2010年第12期2360-2365,共6页Chemical Journal of Chinese Universities

基　　金：国家科技支撑计划项目(批准号:2008BAK45B04);国家质检公益专项(批准号:AHY0709)资助

摘　　要：建立了基于磷元素测量的高分辨电感耦合等离子体质谱(HR-ICP-MS)定量荧光标记DNA的分析方法,该方法测量结果可以溯源到国际基本单位(SI).采用柱层析、超速离心及透析等技术对样品进行纯化,采用芯片电泳和电导率测试仪对其纯度进行了检验.采用优化的微波消解方法对荧光标记DNA样品进行了消解处理.从射频功率、等离子气流速、辅助气流速、雾化气流速、采样深度及获取时间等方面对HR-ICP-MS测量条件进行了优化;从物理性干扰、内标、同位素及元素形态等方面对测量进行了校正.采用优化后的方法对荧光标记DNA样品进行了定量测量,从方法的精密度、标准物质、称量及样品稀释等方面评定了测量结果的不确定度,表明测量结果的扩展不确定度为8.4%(k=2).该方法可用于核酸含量标准物质的定量分析.An accurate mass determination method for fluorescent dye-labeled DNA by high resolution inductively coupled plasma mass spectrometry（HR-ICP-MS） was established.This method allow for the mass of fluorescent dye-labeled DNA to be traceable to the International system of units.The fluorescent dye-labeled DNA was processed by co-lumn chromatography,ultracentrifugation and dialysis.The purity was detected by electrophoresis chip and electrical conductivity.The fluorescent dye-labeled DNA was digested with microwave in nitric acid.The phosphorus from the fluorescent dye-labeled DNA was quantitatively determined by HR-ICP-MS.The parameters of HR-ICP-MS and microwave digestion were optimized.The interference factors of HR-ICP-MS such as physical interference,signal drift interference,isotopes and phosphorus element speciation interference were analyzed and corrected with ultra-filtration,internal standard of Y（88.906）,certified reference material of phosphate,nucleotide and oligo DNA.The uncertainty in measurement was evaluated mainly from method precision,phosphorus certified reference material（CRM）,nitric acid digestion,mass of sample and CRM.The uncertainty（≤8.4%） was smaller than ultraviolet spectrophotometry,fluorescence spectrophotometry and real time PCR.This method will aide in the development of certified reference materials（CRMs） with certified nucleic acid（DNA,RNA） content.

关 键 词：荧光标记DNA 磷 高分辨电感耦合等离子发射质谱 定量 不确定度 

分 类 号：O657.6[理学—分析化学]