Coulomb-type DNA Biosensor Based On Gold Nanoparticles Signal Amplification

With the deeper research on the strueture and furetion of human gene, the study on DNA diagnosis and gene treatment of human disease has been greatly promoted. DNA separation and analysis has taken a more important role in the areas of clinical diagnosis, epidemic prevention, bioengineering and environmental protection. Many new biological technologies emerged and found their applications in these fields. Among them, DNA biosensors are rapidly developed and have received considerable attentions. DNA electrochemical biosensor is a novel and developing technique that combining biochemical, electrochemical, medical and electronic techniques with the advantages of being simple, cheap, reliable, sensitive and selective for genetic detection, and has been a hot topic in the field of biochemistry and medicine.Nanoparticles have caught more and more attention because of their small sizes, large specific surface and good biological compatibility characteristics. Based on their unique physical and electrochemical properties, two kinds of electrochemical DNA biosensors with high performance fabricated by using gold nanoparticles (AuNPs) were achieved. This paper is consistsed of the following three chapters:Chapters1:IntroductionFistly, we introduce the DNA biosensor, including its principle and classification of DNA biosensor. Among these, we emphatically review the principle, progress, the application and development trends of electrochemical DNA biosensors. Secondly, the application of gold nanoparticles on DNA biosensors was introduced. Finally, the background, research ideas, purpose and contents of this paper are described.Chapters2:Selective DNA Detection at Zeptomole Level Based on Coulometric Measurement of Gold Nanoparticle-Mediated Electron Transfer across a Self-Assembled MonolayerA selective DNA sensing with zeptomole detection level is developed based on coulometric measurement of gold nanoparticle (AuNPs)-mediated electron transfer (ET) across a self-assembled monolayer on the gold electrode. After immobilization of a thiolated hairpin-structured DNA probe, an alkanethiol monolayer was self-assembled on the resultant electrode to block [Fe(CN)6]3-/4-in a solution from accessing the electrode. In the presence of DNA target, hybridization between the DNA probe and the DNA target breaks the stem duplex of DNA probe. Consequently, stem moiety at the3’-end of the DNA probes was removed from the electrode surface and made available for hybridization with the reporter DNA-AuNPs conjugates (reporter DNA-AuNPs). The thiolated reporter DNA matches the stem moiety at the3’-end of the DNA probe. AuNPs were then enlarged by immersing the electrode in a growth solution containing HAuCl4and H2O2after the reporter DNA-AuNPs bound onto the electrode surface. The enlarged AuNPs on the electrode restored the ET between the electrode and the [Fe(CN)6]3-/4-, as a result, amplified signals were achieved for DNA target detection using the coulometric measurement of Fe(CN)63-electro-reduction by prolonging the electrolysis time. The quantities of ET on the DNA sensor increased with the increase in DNA target concentration through a linear range of3.0fM to1.0pM when electrolysis time was set to300s, and detection limit was1.0fM. Correspondingly, thousands of DNA (zeptomole) copies were detected in10-μL samples. Furthermore, the DNA sensor showed excellent differentiation ability for single-base mismatch.Chapters3:Highly sensitive detection of DNA using an electrochemical DNA sensor with thionine-capped DNA/gold nanoparticle conjugates as signal tagsA highly sensitive electrochemical DNA (E-DNA) sensor based on the voltammetric detection of thionine, which was capped on diblock DNA/gold nanoparticle (DiDNA/AuNP) conjugates, on a gold electrode was developed. The E-DNA sensor was prepared through the self-assembly of a3’-end thiolated probe DNA (PDNA) on a gold electrode followed by the hybridization of thionine-capped DiDNA/AuNP conjugates to the5’-end of the PDNA. The thionine-capped DiDNA/AuNP conjugate was acted as a tag. In the absence of the target DNA, the flexible single-stranded PDNA supports an efficient contact between tag and electrode, ensuring high current of the E-DNA sensor. Upon hybridization, a rigid probe-target duplex is formed, which pushes the tag away from the electrode and increases the distance between the tag and the electrode, thereby decreasing the current of the E-DNA sensor. The analytical method based on this concept is highly sensitive because the thionine-capped DiDNA/AuNP conjugate contains a large number of electroactive thionine molecules. Under optimal conditions, the dynamic detection range of the target DNA was from0.5pM to50pM, and the detection limit was0.3pM. The DNA sensor also exhibited selectivity against single-base mismatched DNA.