title: Electrochemical DNA Biosensor Based on a Tetrahedral Nanostructure Probe for the Detection of Avian Influenza A (H7N9) Virus.
abstract: A DNA tetrahedral nanostructure-based electrochemical biosensor was developed to detect avian influenza A (H7N9) virus through recognizing a fragment of the hemagglutinin gene sequence. The DNA tetrahedral probe was immobilized onto a gold electrode surface based on self-assembly between three thiolated nucleotide sequences and a longer nucleotide sequence containing complementary DNA to hybridize with the target single-stranded (ss)DNA. The captured target sequence was hybridized with a biotinylated-ssDNA oligonucleotide as a detection probe, and then avidin-horseradish peroxidase was introduced to produce an amperometric signal through the interaction with 3,3',5,5'-tetramethylbenzidine substrate. The target ssDNA was obtained by asymmetric polymerase chain reaction (PCR) of the cDNA template, reversely transcribed from the viral lysate of influenza A (H7N9) virus in throat swabs. The results showed that this electrochemical biosensor could specifically recognize the target DNA fragment of influenza A (H7N9) virus from other types of influenza viruses, such as influenza A (H1N1) and (H3N2) viruses, and even from single-base mismatches of oligonucleotides. Its detection limit could reach a magnitude of 100 fM for target nucleotide sequences. Moreover, the cycle number of the asymmetric PCR could be reduced below three with the electrochemical biosensor still distinguishing the target sequence from the negative control. To the best of our knowledge, this is the first report of the detection of target DNA from clinical samples using a tetrahedral DNA probe functionalized electrochemical biosensor. It displays that the DNA tetrahedra has a great potential application as a probe of the electrochemical biosensor to detect avian influenza A (H7N9) virus and other pathogens at the gene level, which will potentially aid the prevention and control of the disease caused by such pathogens.
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.|‡Institute of Applied Mechanics and Biomedical Engineering, Taiyuan University of Technology, Taiyuan, Shanxi 030024, China.
authors.forename: Shibiao
authors.initials: S
authors.lastname: Dong
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Rongtao
authors.initials: R
authors.lastname: Zhao
authors.affiliation: ∥Clinical Diagnostic Center, 302 Hospital of PLA, Beijing 100039, China.
authors.forename: Jiangong
authors.initials: J
authors.lastname: Zhu
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Lu
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Li
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Shaofu
authors.initials: S
authors.lastname: Qiu
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Leili
authors.initials: L
authors.lastname: Jia
authors.affiliation: ‡Institute of Applied Mechanics and Biomedical Engineering, Taiyuan University of Technology, Taiyuan, Shanxi 030024, China.
authors.forename: Xiong
authors.initials: X
authors.lastname: Jiao
authors.affiliation: §Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, China.
authors.forename: Shiping
authors.initials: S
authors.lastname: Song
authors.affiliation: §Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, China.
authors.forename: Chunhai
authors.initials: C
authors.lastname: Fan
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: RongZhang
authors.initials: R
authors.lastname: Hao
authors.affiliation: †Institute for Disease Control and Prevention, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: HongBin
authors.initials: H
authors.lastname: Song
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D001717:Birds / Q000821:virology*; D019187:Cadmium Compounds / Q000737:chemistry; D003102:Colloids / Q000737:chemistry; D003460:Crystallization; D015342:DNA Probes / Q000737:chemistry*; D055664:Electrochemical Techniques / Q000379:methods*; D064766:Influenza A Virus, H7N9 Subtype / Q000302:isolation & purification*; D005585:Influenza in Birds / Q000821:virology*; D007840:Latex / Q000737:chemistry; D008863:Microspheres; D049329:Nanostructures / Q000737:chemistry*; D017785:Photons; D011109:Polymethacrylic Acids / Q000737:chemistry; D011137:Polystyrenes / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared; D013440:Sulfides / Q000737:chemistry
chemical_list: D019187:Cadmium Compounds; D003102:Colloids; D015342:DNA Probes; D007840:Latex; D011109:Polymethacrylic Acids; D011137:Polystyrenes; D013440:Sulfides; C034939:cadmium sulfide; C030613:polymethacrylic acid
keywords: asymmetric PCR; avian influenza A (H7N9) virus; electrochemical DNA biosensor; pathogen detection; tetrahedral nanostructure probe