title: High yield expression and purification of Chikungunya virus E2 recombinant protein and its evaluation for serodiagnosis.
abstract: Disease caused by Chikungunya virus (CHIKV) is clinically characterized by sudden-onset of fever and severe arthralgia, which may persist for weeks, months, or years after acute phase of the infection. CHIKV is spreading globally; in India it first appeared in the 1960s followed by a quiescent period and then a full-blown remergence in 2006 and sporadic persistence since then. Despite a large number of commercially available diagnostic kits for CHIKV, clinical preparedness and diagnostics suffer from sub-optimal assays. An international diagnostic laboratory survey suggested that there is a critical need for improved CHIKV diagnostics especially in the early acute phase of illness. With the recent studies indicating that a vast majority of human humoral response in CHIKV infection is directed against E2 protein, this supports strong interest to develop CHIKV E2 based serological tests. However, methods to produce large amounts of CHIKV protein are limited. Here we report cloning, expression and purification methods for obtaining a truncated 37kDa Chikungunya E2 protein at a high yield of 65-70mg/l. We found that this purified protein can be reliably used in ELISA and western blot to detect CHIKV specific antibodies in sera from patients who were PCR or IgM positive. Thus, using this protocol, laboratories can make large quantities of purified protein that can be potentially used in CHIKV serological analysis.
authors.affiliation: Department of Pediatrics, All India Institute of Medical Sciences (AIIMS), Ansari Nagar, New Delhi 110029, India.
authors.forename: Anil
authors.initials: A
authors.lastname: Verma
authors.affiliation: ICGEB-Emory Vaccine Center, International Center for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Anmol
authors.initials: A
authors.lastname: Chandele
authors.affiliation: ICGEB-Emory Vaccine Center, International Center for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Kaustuv
authors.initials: K
authors.lastname: Nayak
authors.affiliation: ICGEB-Emory Vaccine Center, International Center for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Murali Krishna
authors.initials: MK
authors.lastname: Kaja
authors.affiliation: Structural Biology & Computational Biology, International Center for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Arockiasamy
authors.initials: A
authors.lastname: Arulandu
authors.affiliation: Department of Pediatrics, All India Institute of Medical Sciences (AIIMS), Ansari Nagar, New Delhi 110029, India.
authors.forename: Rakesh
authors.initials: R
authors.lastname: Lodha
authors.affiliation: Department of Biotechnology, Faculty of Science, Jamia Hamdard, Hamdard Nagar, New Delhi 110062, India. Electronic address: pratimaray.aiims@gmail.com.
authors.forename: Pratima
authors.initials: P
authors.lastname: Ray
mesh_terms: D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral; D065632:Chikungunya Fever / Q000175:diagnosis* / Q000276:immunology; D002646:Chikungunya virus / Q000737:chemistry* / Q000276:immunology; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007075:Immunoglobulin M / Q000097:blood; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins
keywords: Chikungunya; Diagnostics; ELISA; Envelope 2 protein