title: Dengue virus envelope domain III protein based on a tetravalent antigen secreted from insect cells: Potential use for serological diagnosis.
abstract: In the present study, we developed a tetravalent protein by connecting the receptor-binding envelope domain III (EDIII) of the four dengue virus serotypes in the order of D1-D3-D4-D2. Using a baculovirus expression system, the protein was secreted into the supernatant of infected sf9 cells in a stable form with preserved native conformation. Using immobilized affinity chromatography, the recombinant EDIII (rEDIII) protein was purified with a yield of 300μg per 10(6) cells. The purity and reactivity of the protein were determined via SDS-PAGE and Western blot respectively. A MAC-ELISA method based on the secreted rEDIII protein was subsequently established and evaluated using a panel of pre-characterized dengue IgM-positive and -negative human sera. We obtained a specificity of 100% and sensitivity of 93% using this method. Our data collectively suggest that the secreted tetravalent rEDIII protein has potential utility in the diagnosis of dengue virus infections.
authors.affiliation: School of Public Health, WeiFang Medical University, 261053, China; Tianjin International Travel Health Care Center, Tianjin 300456, China.
authors.forename: Guoyu
authors.initials: G
authors.lastname: Niu
authors.affiliation: School of Life Sciences, Tianjin University, Tianjin 300072, China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Pang
authors.affiliation: Tianjin International Travel Health Care Center, Tianjin 300456, China.
authors.forename: Chun
authors.initials: C
authors.lastname: Guan
authors.affiliation: Tianjin International Travel Health Care Center, Tianjin 300456, China. Electronic address: qij@tjciq.gov.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Qi
authors.affiliation: National Institute for Viral Disease Control and Prevention, China CDC, 155 Chang Bai Road, Chang Ping District, Beijing 102206, China. Electronic address: dexinlicdc@gmail.com.
authors.forename: Dexin
authors.initials: D
authors.lastname: Li
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000235:genetics / Q000302:isolation & purification*; D016367:Baculoviridae; D003715:Dengue / Q000175:diagnosis*; D003716:Dengue Virus / Q000235:genetics / Q000302:isolation & purification*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods*; D061987:Sf9 Cells; D014759:Viral Envelope Proteins / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007075:Immunoglobulin M; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins
keywords: Dengue virus; Diagnosis; Secretion; Tetravalent