Expression and purification of recombinant H5HA1 protein of H5N1 avian influenza virus in E. coli and its application in indirect ELISA. Issue 4 (3rd July 2016)

• Record Type: Journal Article
• Authors: Redda, Y. T.; Venkatesh, G.; Kalaiyarasu, S.; Bhatia, S.; Kumar, D. Senthil; Nagarajan, S.; Pillai, A.; Tripathi, S.; Kulkarni, D. D.; Dubey, S. C.
• Is Part Of: Journal of immunoassay & immunochemistry. Volume 37:Issue 4(2016)
• Page Start: 346
• Page End: 358
• Publication Date: 2016-07-03
• Subjects: AIV H5N1 -- haemagglutinin -- expression -- characterization -- indirect ELISA; Immunoassay -- Periodicals; Immunochemistry -- Periodicals; Immunoassay -- periodicals; Immunochemistry -- periodicals; 571.96405
• DOI: 10.1080/15321819.2015.1135160
• Languages: English
• ISSNs: 1532-1819

Abstract

ABSTRACT: The PCR amplified HA1 fragment of H5N1 (H5HA1) avian influenza virus (AIV) hemagglutinin gene was cloned into pET28a (+) expression vector and expressed in Rosetta Blue ( DE3 ) pLys S cells. The recombinant H5HA1 (rH5HA1) protein purified by passive gel elution after SDS-PAGE of the inclusion bodies reacted specifically with H5N1 serum in Western blot analysis. A subtype specific indirect enzyme linked immunosorbent assay (iELISA) using the rH5HA1 protein as the coating antigen was developed for detecting antibodies to H5 subtype of AIV. The assay had 89.04% sensitivity and 95.95% specificity when compared with haemagglutination inhibition test. The Kappa value of 0.842 indicated a perfect agreement between the tests. The iELISA developed can be used for serosurveillance of avian influenza in chickens.