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. 2010 Dec 21;25(6):390–400. doi: 10.1007/s12250-010-3160-y

Detection of bluetongue virus group-specific antigen using monoclonal antibody based sandwich ELISA

Pradeep Narayan Gandhale 1, Veerakyathappa Bhanuprakash 1,✉, Vinayagamurthy Balamurugan 2, Madhusudhan Hosamani 3, Gnanavel Venkatesan 1, Raj Kumar Singh 4
PMCID: PMC8227905  PMID: 21221917

Abstract

A monoclonal antibody (MAb) specific for the bluetongue virus (BTV) group specific antigen (VP7) was characterized for its reactivity with purified virus and recombinant BTV VP7 (rVP7) protein and its suitability for use in the sandwich ELISA. The MAb, designated as 5B5 was specific to VP7 and belongs to IgG2a subclass and was selected for the development of the sELISA in this study. The MAb had a titer of 1:25 with BTV and 1:2 with the rVP7 protein. The sELISA is based on capturing of BTV antigen with VP7 specific MAb followed by detection using BTV polyclonal antiserum raised in rabbits. The assay was evaluated with six cell culture adapted serotypes of BTV that have been isolated from India, 1, 2, 15, 17, 18 and 23. The assay could detect BTV antigen as early as day 8 in blood. It was also successfully applied for the detection of BTV group specific antigen in clinical samples of blood, washed RBCs, buffy coat and plasma. A total of 102 field samples from animals, suspected of being infected with BTV, were tested and 29.42% were positive. The blood samples were also amplified in cell culture which improved the sensitivity of the assay. Results confirmed that the sELISA is rapid and specific.

Key words: Bluetongue, Diagnosis, ELISA, Monoclonal antibody, Sandwich ELISA

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