Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus

doi:10.1016/S2095-3119(13)60332-9

Journal of Integrative Agriculture ›› 2013, Vol. 12 ›› Issue (9): 1638-1643.DOI: 10.1016/S2095-3119(13)60332-9

Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus

NIU Hui-min, HUANG Xin-mei, HAN Kai-kai, LIU Yu-zhuo, ZHAO Dong-min, ZHANG Jing-feng, LIU Fei, LI Tong-tong, ZHOU Xiao-bo, LI Xiang-rui , LI Yin

1.Institute of Veterinary Medicine, Jiangsu Academy of Agricutural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture/National Center for Engineering Research of Veterinary Bio-Products, Nanjing 210014, P.R.China
2.College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, P.R.China

收稿日期:2012-07-10 出版日期:2013-09-01 发布日期:2013-09-15
通讯作者: Correspondence LI Yin, Tel: +86-25-84391687, Fax: +86-25-84390330, E-mail: muziyinyin09@yahoo.com.cn
作者简介:NIU Hui-min, E-mail: hankk0917@126.com
基金资助:
The project was supported by the National Natural Science Foundation of China (31172345) and the Jiangsu Provincial Agricultural Science and Technology Innovation Foundation, China (cx (11)4039).

Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus

NIU Hui-min, HUANG Xin-mei, HAN Kai-kai, LIU Yu-zhuo, ZHAO Dong-min, ZHANG Jing-feng, LIU Fei, LI Tong-tong, ZHOU Xiao-bo, LI Xiang-rui , LI Yin

1.Institute of Veterinary Medicine, Jiangsu Academy of Agricutural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture/National Center for Engineering Research of Veterinary Bio-Products, Nanjing 210014, P.R.China
2.College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, P.R.China

Received:2012-07-10 Online:2013-09-01 Published:2013-09-15
Contact: Correspondence LI Yin, Tel: +86-25-84391687, Fax: +86-25-84390330, E-mail: muziyinyin09@yahoo.com.cn
About author:NIU Hui-min, E-mail: hankk0917@126.com
Supported by:
The project was supported by the National Natural Science Foundation of China (31172345) and the Jiangsu Provincial Agricultural Science and Technology Innovation Foundation, China (cx (11)4039).

摘要/Abstract

摘要： In order to establish double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for detection of duck or goose flavivirus, polyclonal antibody against the flavivirus strain JS804 in geese and monoclonal antibody against the E protein of flavivirus strain JS804 in geese were used as the capture antibody and detection antibody, respectively. The optimal dilution of the capture antibody and detecting antibody capable of detecting the flavivirus strain JS804 in geese were 1:3 200 and 1:160 in the check-board titration, respectively. The reaction time of sample was 1 h, and the optimal working dilution of HRP-labeled goat-anti-mouse IgG was 1:10 000. The positive standard value was 0.247 (OD450 nm). The geese flavivirus could be detected at a minimal concentration of 1.875 μg mL-1. The ELISA had no cross-reaction with Newcastle disease virus (NDV), Avian influenza virus (AIV), Infectious bronchitis virus (IBV), Infectious bursal disease virus (IBDV), Duck hepatitis virus (DHV), and Gosling plague virus (GPV). Twenty clinical samples were detected by the DAS-ELISA and RT-PCR respectively, with the agreement rate of 75%. The results revealed that the DAS-ELISA possessed favorable specificity and higher sensitivity, indicating a suitable method for rapid detection of the duck or goose flavivirus.

关键词: goose , flavivirus , double antibody sandwich ELISA , monoclonal antibodies

Abstract: In order to establish double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for detection of duck or goose flavivirus, polyclonal antibody against the flavivirus strain JS804 in geese and monoclonal antibody against the E protein of flavivirus strain JS804 in geese were used as the capture antibody and detection antibody, respectively. The optimal dilution of the capture antibody and detecting antibody capable of detecting the flavivirus strain JS804 in geese were 1:3 200 and 1:160 in the check-board titration, respectively. The reaction time of sample was 1 h, and the optimal working dilution of HRP-labeled goat-anti-mouse IgG was 1:10 000. The positive standard value was 0.247 (OD450 nm). The geese flavivirus could be detected at a minimal concentration of 1.875 μg mL-1. The ELISA had no cross-reaction with Newcastle disease virus (NDV), Avian influenza virus (AIV), Infectious bronchitis virus (IBV), Infectious bursal disease virus (IBDV), Duck hepatitis virus (DHV), and Gosling plague virus (GPV). Twenty clinical samples were detected by the DAS-ELISA and RT-PCR respectively, with the agreement rate of 75%. The results revealed that the DAS-ELISA possessed favorable specificity and higher sensitivity, indicating a suitable method for rapid detection of the duck or goose flavivirus.

Key words: goose , flavivirus , double antibody sandwich ELISA , monoclonal antibodies

NIU Hui-min, HUANG Xin-mei, HAN Kai-kai, LIU Yu-zhuo, ZHAO Dong-min, ZHANG Jing-feng, LIU Fei, LI Tong-tong, ZHOU Xiao-bo, LI Xiang-rui , LI Yin. Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus[J]. Journal of Integrative Agriculture, 2013, 12(9): 1638-1643.

参考文献

[1]Hou M R, Hou X L, Gao J F, Liu Z G, Wang J, Yang M F.2011. Development of a double antibody sandwichELISA for detection of Bovine rotavirus. Journal ofHeilongjiang Bayi Agricultural University, 3, 19-23 (in Chinese)

[2]Huang L 2004. The Theory of Statistics. China StatisticsPress, Beijing. (in Chinese)

[3]Huang X M, Li Y, Zhao D M, Liu Y Z, Zhang J F, He K W,Hou J B, Niu H M, Li B, Wen L B, et al. 2011. Isolationand identification of a novel flavivirus strain JS804 ingeese. Jiangsu Journal of Agricultural Science, 2, 354-360 (in Chinese)

[4]Ji Y Y, Guo W, Wang X J, Wang Z, Lu G, Zhao L P, Li H M,Xiang W H. 2011. Development of double antibodysandwich ELISA for detection of Equine influenza virus.Chinese Journal of Animal and Veterinary Sciences,42, 679-684 (in Chinese)

[5]Niu N N. 2010. Development of McAbs against H9 AIVand establishment of antigen capture ELISA fordetecting H9N2 avian influenza virus. MSc thesis,Nanjing Agricultural University, Nanjing. pp. 27-29 (in Chinese)

[6]Su J L, Li S, Hu X D, Yu X L, Wang Y Y, Liu P P, Lu X S,Zhang G Z, Hu X M, Liu D, et al. 2011. Duck egg-dropsyndrome caused by BYD virus, a new tembusu-relatedflavivirus. PLoS ONE, 6, e18106.

[7]Tang Y, Diao Y, Yu C, Gao X, Chen L, Zhang D. 2011. Rapiddetection of Tembusu virus by reverse-transcriptionloop-mediated isothermal amplification (RT-LAMP).Transboundary and Emerging Diseases, doi: 10.1111/j.1865-1682

[8]2011.01257.xTeng Q Y, Yan P X, Zhang X, Yan L P, Li Z J. 2010. A novelFlavivirus causing duck egg drops and death. ChineseJournal of Animal Infectious Diseases, 18, 1-4

[9]Wan C H, Shi S H, Cheng L F, Chen H M, Fu G H, Peng C X,Liu F, Lin J S, Huang Y. 2011. Establishment of RT-PCRfor detecting duck hemorrhagic ovaritis causing abruptegg-laying reduction in ducks. Fujian Journal ofAgricultural Sciences, 26, 10-12 (in Chinese)

[10]Wang J K, Cheng S P, Yi L, Luo G L, Luo B, Wu W, Zhao CF. 2011. Establishment of double antibody sandwichELISA for detection of Mink enteritis virus. ChineseVeterinary Science, 41, 183-187 (in Chinese)

[11]Wang Y L, Yuan X Y, Li Y F, Yu K X, Yang J X, Xu H Y,Zhang Y X, Yu K Z, Liao M, Qin Z M. 2011. Rapiddetection of newly isolated Tembusu-related Flavivirusby reverse-transcription loop-mediated isothermalamplification assay. Virology Journal, 8, 553-560

[12]Yan L P, Yan P X, Zhou J W, Teng Q Y, Li Z J. 2011.Establishing a TaqMan-based real-time PCR assay forthe rapid detection and quantification of the newlyemerged duck Tembusu virus. Virology Journal, 8, 464-471

[13]Yan P X, Li G J, Wu X G, Yan L P, Teng Q Y, Li Z J. 2011.Rapid identification of duck Tembusu virus by thenested RT-PCR. Chinese Journal of Animal InfectiousDiseases, 19, 34-37

[14]Zhu B. 2009. Development and application of MonoclonalAntibodies against classical swine fever virus. MScthesis, Huazhong Agricultural University, Wuhan. p.20. (in Chinese)

Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus

Development of Double Antibody Sandwich ELISA for Detection of Duck or Goose Flavivirus

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 2

编辑推荐

Metrics

[1]	MA Xiao-yan, WU Han-wen, JIANG Wei-li, MA Ya-jie, MA Yan. Goosegrass (Eleusine indica) density effects on cotton (Gossypium hirsutum)[J]. Journal of Integrative Agriculture, 2015, 14(9): 1778-1785.
[2]	CHEN Jing-chao, HUANG Hong-juan, WEI Shou-hui, ZHANG Chao-xian, HUANG Zhao-feng. Characterization of glyphosate-resistant goosegrass (Eleusine indica) populations in China[J]. Journal of Integrative Agriculture, 2015, 14(5): 919-925.