title: An electrochemical aptasensor for detection of IFN-γ using graphene and a dual signal amplification strategy based on the exonuclease-mediated surface-initiated enzymatic polymerization.
abstract: Tuberculosis is one of the major health problems in the world. The cytokine interferon γ (IFN-γ) is associated with the disease-specific immune responses and is used as a tuberculosis diagnosis marker. In this study, a novel electrochemical aptasensor was developed for IFN-γ detection based on the exonuclease-catalyzed target recycling and the TdT-mediated cascade signal amplification. To construct the aptasensor, a previously hybridized double-stranded DNA (capture probe hybridization with a complementary IFN-γ binding aptamer) was immobilized on a gold nanoparticle-graphene (Au-Gra) nanohybrid film-modified electrode. In the presence of IFN-γ, the formation of an aptamer-IFN-γ complex leads to the liberation of the aptamer from the double-stranded DNA (dsDNA). Using exonuclease, the aptamer was selectively digested, and IFN-γ was released for the target recycling. A large amount of single-stranded capture probes formed and led to the hybridization with signal probe-labelled Au@Fe3O4. Then, the labelled signal probe sequences were catalyzed at the 3'-OH group by terminal deoxynucleotidyl transferase (TdT) to form a long single-stranded DNA structure. As a result, the electron mediator hexaammineruthenium(III) chloride ([Ru(NH3)6](3+)) electrostatically adsorbed onto DNA producing a strong electrochemical signal which can be used to quantitatively measure the IFN-γ levels. With the conducting nanomaterial Au-Gra as a substrate and the target recycling-based surface-initiated enzymatic polymerization-mediated signal amplification strategy, the proposed aptasensor displayed a broad linearity with a low detection limit of 0.003 ng mL(-1). Moreover, the resulting aptasensor exhibited good specificity, acceptable reproducibility and stability, which makes this method versatile and suitable for detecting IFN-γ and other biomolecules.
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Chang
authors.initials: C
authors.lastname: Liu
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Guiming
authors.initials: G
authors.lastname: Xiang
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Dongneng
authors.initials: D
authors.lastname: Jiang
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Linlin
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Fei
authors.initials: F
authors.lastname: Liu
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Fukang
authors.initials: F
authors.lastname: Luo
authors.affiliation: Department of Clinical Laboratory, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, P. R. China. Luofukang@tmmu.edu.cn xqyyjyk@foxmail.com.
authors.forename: Xiaoyun
authors.initials: X
authors.lastname: Pu
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000378:metabolism*; D015374:Biosensing Techniques / Q000379:methods*; D055664:Electrochemical Techniques / Q000379:methods; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D006046:Gold / Q000737:chemistry*; D006108:Graphite / Q000737:chemistry*; D006801:Humans; D007371:Interferon-gamma / Q000032:analysis / Q000097:blood* / Q000378:metabolism; D057230:Limit of Detection; D049329:Nanostructures / Q000737:chemistry / Q000648:ultrastructure; D058105:Polymerization; D015203:Reproducibility of Results
chemical_list: D052157:Aptamers, Nucleotide; D006046:Gold; D006108:Graphite; D007371:Interferon-gamma; D005090:Exodeoxyribonucleases