Publication: Production of monoclonal antibodies specific to Macrobrachium rosenbergii nodavirus using recombinant capsid protein
Issued Date
2012-03-20
Resource Type
ISSN
16161580
01775103
01775103
DOI
Other identifier(s)
2-s2.0-84861668619
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Mahidol University
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SCOPUS
Bibliographic Citation
Diseases of Aquatic Organisms. Vol.98, No.2 (2012), 121-131
Suggested Citation
Pradit Wangman, Saengchan Senapin, Parin Chaivisuthangkura, Siwaporn Longyant, Sombat Rukpratanporn, Paisarn Sithigorngul Production of monoclonal antibodies specific to Macrobrachium rosenbergii nodavirus using recombinant capsid protein. Diseases of Aquatic Organisms. Vol.98, No.2 (2012), 121-131. doi:10.3354/dao02431 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/13478
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Title
Production of monoclonal antibodies specific to Macrobrachium rosenbergii nodavirus using recombinant capsid protein
Abstract
The gene encoding the capsid protein of Macrobrachium rosenbergii nodavirus (MrNV) was cloned into pGEX-6P-1 expression vector and then transformed into the Escherichia coli strain BL21. After induction, capsid protein-glutathione-S- transferase (GST-MrNV; 64 kDa) was produced. The recombinant protein was separated using SDS-PAGE, excised from the gel, electro-eluted and then used for immunization for monoclonal antibody (MAb) production. Four MAbs specific to the capsid protein were selected and could be used to detect natural MrNV infections in M. rosenbergii by dot blotting, Western blotting and immunohistochemistry without cross-reaction with uninfected shrimp tissues or other common shrimp viruses. The detection sensitivity of the MAbs was 10 fmol μl-1 of the GST-MrNV, as determined using dot blotting. However, the sensitivity of the MAb on dot blotting with homogenate from naturally infected M. rosenbergii was approximately 200-fold lower than that of 1-step RT-PCR. Immunohistochemical analysis using these MAbs with infected shrimp tissues demonstrated staining in the muscles, nerve cord, gill, heart, loose connective tissue and inter-tubular tissue of the hepatopancreas. Although the positive reactions occurred in small focal areas, the immunoreactivity was clearly demonstrated. The MAbs targeted different epitopes of the capsid protein and will be used to develop a simple immunoassay strip test for rapid detection of MrNV. © Inter-Research 2012.