title: Expression and purification of recombinant H5HA1 protein of H5N1 avian influenza virus in E. coli and its application in indirect ELISA.
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) pLysS 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.
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: Y T
authors.initials: YT
authors.lastname: Redda
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: G
authors.initials: G
authors.lastname: Venkatesh
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: S
authors.initials: S
authors.lastname: Kalaiyarasu
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: S
authors.initials: S
authors.lastname: Bhatia
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: D Senthil
authors.initials: DS
authors.lastname: Kumar
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: S
authors.initials: S
authors.lastname: Nagarajan
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: A
authors.initials: A
authors.lastname: Pillai
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: S
authors.initials: S
authors.lastname: Tripathi
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: D D
authors.initials: DD
authors.lastname: Kulkarni
authors.affiliation: a ICAR - National Institute of High Security Animal Diseases , Anand Nagar, Bhopal , Madhya Pradesh , India.
authors.forename: S C
authors.initials: SC
authors.lastname: Dubey
mesh_terms: D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics* / Q000276:immunology; D006385:Hemagglutination Inhibition Tests; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics* / Q000276:immunology; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D014764:Viral Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*
chemical_list: D011994:Recombinant Proteins; D014764:Viral Proteins
keywords: AIV H5N1; characterization; expression; haemagglutinin; indirect ELISA