基于末端脱氧核苷酸转移酶扩增DNA-铜纳米簇荧光传感检测L-组氨酸  

Terminal Deoxynucleotidyl Transferase Amplification Based DNA-Copper Nanoclusters Sensor for Detection of L-Histidine

作　　者：肖慧 何婧琳 肖昊[2] 杨婵 冯泽猛[2] 印遇龙[2] 曹忠 

机构地区：[1]长沙理工大学化学与生物工程学院,电力与交通材料保护湖南省重点实验室,微纳生物传感与食品安全检测协同创新中心,长沙410114 [2]中国科学院亚热带农业生态研究所,长沙410125

出　　处：《分析化学》2017年第10期1517-1522,共6页Chinese Journal of Analytical Chemistry

基　　金：国家自然科学基金项目(Nos.31527803,21545010,21645009);中国科学院科技服务网络STS计划(No.KFJ-SW-STS-173)项目资助~~

摘　　要：利用末端脱氧核苷酸转移酶(TdT)扩增形成聚胸腺嘧啶(T)DNA模板,制备了聚T铜纳米簇(TS-CuNCs),构建了一种用于L-组氨酸(L-His)检测的荧光传感分析新方法。TdT酶在d TTP存在下合成聚T单链DNA核苷酸序列。由于胸腺嘧啶和Cu^(2+)之间的亲合力,聚T单链DNA作为合成铜纳米簇(CuNCs)的模板,加入还原剂后形成CuNCs,荧光强度增强。在L-His存在下,L-组氨酸的咪唑基与Cu^(2+)螯合形成L-His-Cu^(2+)配合物,因而进入聚胸腺嘧啶序列中的Cu^(2+)量减少,使得合成的CuNCs数量减少,导致荧光信号减弱。实验结果表明,体系荧光响应信号与L-His浓度的对数值在5.0×10^(-9)~5.0×10^(-4)mol/L范围内呈线性关系,检出限达到3.4×10^(-9)mol/L。本方法用于实际尿液样品中L-组氨酸检测的回收率为97.4%~104.6%,在生物医学及临床诊断中具有潜在应用价值。A terminal deoxynucleotidyl transferase（TdT） amplification based DNA-copper nanoclusters（CuNCs） sensor was developed for detection of L-histidine（L-His）.Single strand DNA containing polythymine（T） sequences were synthesized by TdT in the presence of d TTP.In blank control,poly-T sequences worked as templates of CuNCs due to the affinity between thymine and copper ions（Ⅱ）.Fluorescence intensity was enhanced when CuNCs formed with reducing agents.In the presence of L-His,the imidazolyl group of L-His worked as a chelating agent that formed L-His-Cu^（2＋）chelated complex.Thus less copper ions were induced in poly-T sequences,and less CuNCs were obtained to produce week fluorescence signals.A good linear correlation was obtained between fluorescence change and the logarithm of the L-His concentration over the range of 5.0×10^（-9）-5.0×10^（-4）mol/L.The detection limit was estimated as 3.4×10^（-9）mol/L.And the recoveries were 97.4%-104.6% for the actual urine samples.Compared with other methods of synthetic CuNCs,this method allowed to specifically determining L-histidine without template or labeling,which showed good potential in biomedical and clinical analysis.

关 键 词：末端脱氧核苷酸转移酶 聚胸腺嘧啶 铜纳米簇 L-组氨酸 荧光分析 

分 类 号：O657.3[理学—分析化学] R446[理学—化学]