Scientia Agricultura Sinica ›› 2014, Vol. 47 ›› Issue (5): 1006-1012.doi: 10.3864/j.issn.0578-1752.2014.05.017

• ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT • Previous Articles     Next Articles

Nano-Materials-Based Electrochemical Immunosensor for the Detection of Avian Reovirus

 HUANG  Jiao-Ling, XIE  Zhi-Xun, XIE  Li-Ji, TENG  Li-Qiong, HUANG  Li, LIU  Jia-Bo, PANG  Yao-Shan, DENG  Xian-Wen, XIE  Zhi-Qin, FAN  Qing, LUO  Si-Si   

  1. Guangxi Veterinary Research Institute/Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Nanning 530001
  • Received:2013-05-28 Online:2014-03-01 Published:2013-11-29

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Abstract: 【Objective】A novel assay of nano-materials-based electrochemical immunosensor for the detection of avian reovirus (ARV) was established. 【Method】 Graphene sheet-chitosan homogeneous composite was firstly dispersed in acetic acid solution and then the gold nanoparticles (AuNPs) were in situ synthesized at the composite. A graphene sheet-chitosan-gold nano-particles (G-Chi-AuNPs) modified gold electrode was used as the sensor platform to detect ARV target in samples. Similar method was used to synthesize graphene sheet-chitosan-silver nanoparticles composite (G-Chi-AgNPs), and then avian reovirus monoclonal antibodies(ARV-MAb) was immobilized on the G-Chi-AgNPs composite as the signal amplification materials to construct the immunosensor. This electrochemical immunosensor was used to test avian influenza viruses H5N1, H3N6 and H9N2 subtypes, Newcastle disease virus (NDV), laryngotracheitis virus (LTV), infectious bronchitis virus (IBV) and infectious bursal disease virus (IBDV) to ensure the electrochemical immunosensor’s specificity. The electrochemical immunosensor was used to detect 106.5-100.5 TCID50/mL ARV to ensure the electrochemical immunosensor’s sensitivity. Then this electrochemical immunosensor was used to detect the clinical samples to evaluate the practical use of electrochemical immunosensor.【Result】The optimization of the G-Chi-AgNPs-ARV-MAb composite incubation time was studied in detail, and the optimal condition was 40 min. When the sample is tested positive, ARV and ARV-MAb specific combination, G-Chi-AgNPs-ARV-MAb composite are fixed onto the electrode surface, so it’s linear sweep voltammetry curve shows an AgNPs oxidation peak. When the sample is tested negative, it’s linear sweep voltammetry curve dose not show an AgNPs oxidation peak. The developed immunosensor is specific to ARV and does not react with non-target avian viruses when a test sample consists of avian influenza viruses H5N1, H3N6 and H9N2 subtypes, Newcastle disease virus (NDV), laryngotracheitis virus (LTV), infectious bronchitis virus (IBV) and infectious bursal disease virus (IBDV).The electrochemical immunosensor detected the ARV target at a titer higher than 101.5 TCID50/mL. Twelve specimens of ARV were identified from the 22 clinical samples by electrochemical immunosensor assays developed, which was consistant with that of virus isolation. 【Conclusion】 The novel nano-materials-based electrochemical immunosensor for ARV is highly specific and sensitive, and has great potential for efficient diagnosis and monitoring of ARV in affected poultry.

Key words: graphene sheet , electrochemical immunosensor , gold nano-particles , Ag nano-particles , avian reovirus , avian reovirus monoclonal antibodies

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