Zhou Na, Yang Tao, Jiang Chen, Du Meng, Jiao Kui
Key Laboratory of Eco-chemical Engineering, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao, China.
Talanta. 2009 Jan 15;77(3):1021-6. doi: 10.1016/j.talanta.2008.07.058. Epub 2008 Aug 7.
A polyaniline nanofibers (PAN(nano))/carbon paste electrode (CPE) was prepared via dopping PAN(nano) in the carbon paste. The nanogold (Au(nano)) and carbon nanotubes (CNT) composite nanoparticles were bound on the surface of the PAN(nano)/CPE. The immobilization and hybridization of the DNA probe on the Au(nano)-CNT/PAN(nano) films were investigated with differential pulse voltammetry (DPV) and cyclic voltammetry (CV) using methylene blue (MB) as indicator, and electrochemical impedance spectroscopy (EIS) using Fe(CN)(6) as redox probe. The voltammetric peak currents of MB increased dramatically owing to the immobilization of the probe DNA on the Au(nano)-CNT/PAN(nano) films, and then decreased obviously owing to the hybridization of the DNA probe with the complementary single-stranded DNA (cDNA). The electron transfer resistance (R(et)) of the electrode surface increased after the immobilization of the probe DNA on the Au(nano)-CNT/PAN(nano) films and rose further after the hybridization of the probe DNA. The remarkable difference between the R(et) value at the DNA-immobilized electrode and that at the hybridized electrode could be used for the label-free EIS detection of the target DNA. The loading of the DNA probe on Au(nano)-CNT/PAN(nano) films was greatly enhanced and the sensitivity for the target DNA detection was markedly improved. The sequence-specific DNA of phosphinothricin acetyltransferase (PAT) gene and the polymerase chain reaction (PCR) amplification of nopaline synthase (NOS) gene from transgenically modified beans were determined with this label-free EIS DNA detection method. The dynamic range for detecting the PAT gene sequence was from 1.0 x 10(-12)mol/L to 1.0 x 10(-6)mol/L with a detection limit of 5.6 x 10(-13)mol/L.
通过将聚苯胺纳米纤维（PAN(nano)）掺杂到碳糊中制备了聚苯胺纳米纤维/碳糊电极（CPE）。纳米金（Au(nano)）和碳纳米管（CNT）复合纳米粒子被固定在PAN(nano)/CPE表面。以亚甲基蓝（MB）为指示剂，采用差分脉冲伏安法（DPV）和循环伏安法（CV）研究了DNA探针在Au(nano)-CNT/PAN(nano)膜上的固定化和杂交情况，并以Fe(CN)(6)为氧化还原探针采用电化学阻抗谱（EIS）进行研究。由于探针DNA固定在Au(nano)-CNT/PAN(nano)膜上，MB的伏安峰电流显著增加，随后由于DNA探针与互补单链DNA（cDNA）杂交而明显降低。在Au(nano)-CNT/PAN(nano)膜上固定探针DNA后，电极表面的电子转移电阻（R(et)）增加，探针DNA杂交后进一步升高。DNA固定电极和杂交电极的R(et)值之间的显著差异可用于目标DNA的无标记EIS检测。DNA探针在Au(nano)-CNT/PAN(nano)膜上的负载量大大提高，目标DNA检测的灵敏度显著提高。采用这种无标记EIS DNA检测方法测定了草丁膦乙酰转移酶（PAT）基因的序列特异性DNA以及转基因大豆中胭脂碱合酶（NOS）基因的聚合酶链反应（PCR）扩增产物。检测PAT基因序列的动态范围为1.0×10(-12)mol/L至1.0×10(-6)mol/L，检测限为5.6×10(-13)mol/L。
