title: A sensitive and specific antigen detection assay for Middle East respiratory syndrome coronavirus.
abstract: Since its emergence in 2012, more than 900 laboratory-confirmed cases of Middle East respiratory syndrome (MERS) have been reported with a fatality rate of more than 30%. However, no antigen detection assay for commercial use is available for diagnosis. In this study, the full-length nucleocapsid protein (NP) gene of MERS coronavirus (MERS-CoV) was cloned and expressed in Escherichia coli. A MERS-CoV NP capture enzyme-linked immunosorbent assay (ELISA) using two MERS-CoV-NP-specific monoclonal antibodies (MAbs) generated was developed. The ELISA was evaluated using 129 nasopharyngeal aspirates (NPAs) positive for various respiratory viruses and simulated positive NPAs by adding serial dilutions of MERS-CoV. Using a cutoff OD of 0.19, all 129 NPAs positive for respiratory viruses showed very low OD, with a specificity of 100%. For the two simulated MERS-CoV-positive NPAs with serial dilutions of live MERS-CoV, all samples with ≥10 50% tissue culture infective dose (TCID50)/0.1 mL showed positive results. For the 10 additional NPAs with 20 and 200 TCID50/0.1 mL of live MERS-CoV added, all were positive. A highly sensitive and specific MAbs-based antigen capture ELISA has been developed for MERS. This sensitive and specific antigen capture ELISA should be useful for detection of MERS-CoV in human and dromedaries and in field studies.
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Life Sciences, Xiamen University , Xiamen 361102, Fujian Province, China ; National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University , Xiamen 361102, Fujian Province, China.
authors.forename: Yixin
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China.
authors.forename: Kwok-Hung
authors.initials: KH
authors.lastname: Chan
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Life Sciences, Xiamen University , Xiamen 361102, Fujian Province, China ; National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University , Xiamen 361102, Fujian Province, China.
authors.forename: Yahong
authors.initials: Y
authors.lastname: Kang
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China ; State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong , Hong Kong, China ; Research Centre of Infection and Immunology, The University of Hong Kong , Hong Kong, China ; Carol Yu Centre for Infection, The University of Hong Kong , Hong Kong, China.
authors.forename: Honglin
authors.initials: H
authors.lastname: Chen
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China.
authors.forename: Hayes K H
authors.initials: HK
authors.lastname: Luk
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China.
authors.forename: Rosana W S
authors.initials: RW
authors.lastname: Poon
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China ; State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong , Hong Kong, China ; Research Centre of Infection and Immunology, The University of Hong Kong , Hong Kong, China ; Carol Yu Centre for Infection, The University of Hong Kong , Hong Kong, China.
authors.forename: Jasper F W
authors.initials: JF
authors.lastname: Chan
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China ; State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong , Hong Kong, China ; Research Centre of Infection and Immunology, The University of Hong Kong , Hong Kong, China ; Carol Yu Centre for Infection, The University of Hong Kong , Hong Kong, China.
authors.forename: Kwok-Yung
authors.initials: KY
authors.lastname: Yuen
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Life Sciences, Xiamen University , Xiamen 361102, Fujian Province, China ; National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health, Xiamen University , Xiamen 361102, Fujian Province, China.
authors.forename: Ningshao
authors.initials: N
authors.lastname: Xia
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China ; State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong , Hong Kong, China ; Research Centre of Infection and Immunology, The University of Hong Kong , Hong Kong, China ; Carol Yu Centre for Infection, The University of Hong Kong , Hong Kong, China.
authors.forename: Susanna K P
authors.initials: SK
authors.lastname: Lau
authors.affiliation: Department of Microbiology, The University of Hong Kong , Hong Kong, China ; State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong , Hong Kong, China ; Research Centre of Infection and Immunology, The University of Hong Kong , Hong Kong, China ; Carol Yu Centre for Infection, The University of Hong Kong , Hong Kong, China.
authors.forename: Patrick C Y
authors.initials: PC
authors.lastname: Woo
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D000956:Antigens, Viral / Q000032:analysis*; D002162:Camelus; D018352:Coronavirus Infections / Q000175:diagnosis* / Q000276:immunology; D004197:Disease Reservoirs; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards*; D005260:Female; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D065207:Middle East Respiratory Syndrome Coronavirus / Q000276:immunology* / Q000302:isolation & purification; D019590:Nucleocapsid Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D015047:Zoonoses / Q000175:diagnosis / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D019590:Nucleocapsid Proteins
keywords: ELISA; MERS coronavirus; antigen; detection