DNA sensor by using electrochemiluminescence of acridinium ester initiated by tripropylamine
- 373 Downloads
It was found that tripropylamine (TPA) could be used as a coreactant to initiate the electrochemiluminescence (ECL) of acridinium NHS ester (AE NHS) labels attached to DNA. The radicals generated in the electro-oxidation process of TPA reacted with AE NHS to form the excited N-methylacridone, giving rise to light emission. The AE/TPA ECL system was for the first time used as the detection system for developing an ECL-based DNA sensor. In the protocol, streptavidin-modified gold nanoparticles were firstly immobilized onto a thiol-treated gold electrode. The streptavidin could specifically interact with the biontinylated capture DNA. Afterwards, the target DNA and the AE-labeled report DNA were conjugated onto the electrode step by step due to the hybridization reactions, and a sandwich-type sensor was fabricated. The ECL signals of the sensor were obtained under pulse potential condition in alkaline solution containing 50.0 mmol L−1 TPA. Under optimized experimental conditions, the linear range of the DNA sensor for the determination of the target DNA was from 5.0 × 10−15 to 5.0 × 10−12 mol L−1. The detection limit (S/N = 3) was 3.0 × 10−15 mol L−1. Moreover, the sensor could specifically recognize the target DNA against one base-pair mismatched sequences, two base-pair mismatched sequences, and the noncomplementary sequences. It is of great application potential in clinic analysis.
KeywordsElectrochemiluminescence DNA sensor Acridinium ester Gold nanoparticles
This work was supported by the National Natural Science Foundation of P. R. China (grant nos. 20625517 and 20573101) and the Overseas Outstanding Young Scientist Program of the Chinese Academy of Sciences.
- 2.Colinas RJ, Bellisario R, Pass KA (2000) Multiplexed genotyping of β-Globin variants from PCR-amplified newborn blood spot DNA by hybridization with Allele-specific oligodeoxynucleotides coupled to an array of fluorescent microspheres. Clin Chem 46(7):996–998Google Scholar
- 5.Harriff MJ, Wu M, Kent ML, Bermudez LE (2008) Species of environmental Mycobacteria differ in their abilities to grow in human, mouse, and carp macrophages and with regard to the presence of Mycobacterial virulence genes, as observed by DNA microarray hybridization. Appl Environ Microb 74(1):275–285CrossRefGoogle Scholar
- 20.Romaschin AD, Walker PM (2000) Endotoxin activity in whole blood by neutrophil chemiluminescence—a novel analytical paradigm. Clin Chem 46(9):1504–1506Google Scholar
- 33.Bard A, Debad J, Leland J, Sigal G, Wilbur J, Wohlstadter J (2000) Chemiluminescence, electrogenerated. Encyclopedia of Analytical Chemistry: Applications, Theory and Instrumentation 2:9842–9849Google Scholar
- 35.Arnold LJ Jr, Hammond PW, Wiese WA, Nelson NC (1989) Assay formats involving acridinium-ester-labeled DNA probes. Clin Chem 35(8):1588–1594Google Scholar
- 36.Piran U, Kohn DW, Uretsky LS, Bernier D, Barlow EH, Niswander CA, Stastny M (1987) Immunochemiluminometric assay of creatine kinase MB with a monoclonal antibody to the MB isoenzyme. Clin Chem 33(9):1517–1520Google Scholar
- 37.Weeks I, Beheshti I, McCapra F, Campbell AK, Woodhead JS (1983) Acridinium esters as high-specific-activity labels in immunoassay. Clin Chem 29(8):1474–1479Google Scholar
- 38.Goto M, Oka S, Okuzumi K, Kimura S, Shimada K (1991) Evaluation of acridinium-ester-labeled DNA probes for identification of Mycobacterium tuberculosis and Mycobacterium avium–Mycobacterium intracellular complex in culture. J Clin Microbiol 29(11):2473–2476Google Scholar
- 41.Ou C, McDonough S, Cabanas D, Ryder T, Harper M, Moore J, Schochetman G (1990) Rapid and quantitative detection of enzymatically amplified HIV-1 DNA using chemiluminescent oligonucleotide probes. AIDS Res Hum Retrov 6(11):1323–1329Google Scholar
- 42.Kolbert C, Connolly J, Lee M, Persing D (1995) Detection of the Staphylococcal mecA gene by chemiluminescent DNA hybridization. J Clin Microbiol 33(8):2179–2182Google Scholar
- 49.Wu K, Fei J, Bai W, Hu S (2003) Direct electrochemistry of DNA, guanine and adenine at a nanostructured film-modified electrode. Anal Bioanal Chem 376(2):205–209Google Scholar