Biosensor for multiplex detection of two DNA target sequences using enzyme-functionalized Au nanoparticles as signal amplification.
Multiplex electrochemical detection of two DNA target sequences in one sample using enzyme-functionalized Au nanoparticles (AuNPs) as catalytic labels for was proposed. This DNA sensor was fabricated using a "sandwich" detection strategy, involving two kinds of capture probes DNA immobilized on glassy carbon electrode (GCE), and hybridization with target DNA sequences, which further hybridized with the reporter DNA loaded on the AuNPs. The AuNP contained two kinds of DNA sequences, one was complementary to the target DNA, while the other was noncomplementary to the target. The noncomplementary sequences were linked with horseradish peroxidase (HRP) and alkaline phosphatase (ALP), respectively. Enhanced detection sensitivity was obtained where the AuNPs carriers increased the amount of enzyme molecules per hybridization. Electrochemical signals were generated from the enzymatic products produced from the substrates catalyzed by HRP and ALP. Under optimal conditions, a 33-mer sequence could be quantified over the ranges from 1.5x10(-13) to 5.0x10(-12) M with a detection limit of 1.0x10(-13) M using HRP-AuNP as labels, and a 33-mer sequence could be quantified over the ranges from 4.5x10(-11) M to 1.0x10(-9) M with a detection limit of 1.2x10(-11) M using ALP-AuNP as labels.
Li
Xue-Mei
XM
Key Laboratory of Eco-chemical Engineering, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China.
Fu
Pei-Yu
PY
Liu
Jin-Ming
JM
Zhang
Shu-Sheng
SS
D000469:Alkaline Phosphatase / Q000737:chemistry / Q000378:metabolism; D015374:Biosensing Techniques / Q000379:methods*; D002244:Carbon / Q000737:chemistry; D004247:DNA / Q000032:analysis*; D055664:Electrochemical Techniques / Q000379:methods; D004566:Electrodes; D006046:Gold / Q000737:chemistry*; D006735:Horseradish Peroxidase / Q000737:chemistry / Q000378:metabolism; D053768:Metal Nanoparticles / Q000737:chemistry*; D009693:Nucleic Acid Hybridization / Q000379:methods
D002244:Carbon; D006046:Gold; D004247:DNA; D006735:Horseradish Peroxidase; D000469:Alkaline Phosphatase