title: Evaluation of envelope domain III-based single chimeric tetravalent antigen and monovalent antigen mixtures for the detection of anti-dengue antibodies in human sera.
abstract: BACKGROUND
  Flavivirus cross-reactive antibodies in human sera interfere with the definitive identification of dengue virus (DENV) infections especially in areas with multiple co-circulating flaviviruses. Use of DENV envelope domain-III (EDIII) can partially resolve the problem. This study has examined the effect of (i) incorporating the EDIIIs of four DENV serotypes into a single chimeric antigen, and (ii) immobilizing the antigen through specific interaction on the sensitivity and specificity of anti-DENV antibody detection.
  
  
  METHODS
  A sera panel (n = 164) was assembled and characterized using commercial kits for infection by DENV and a host of other pathogens. Anti-DENV antibodies of both IgM and IgG classes in this panel were detected in indirect ELISAs using a mixture of monovalent EDIIIs, a chimeric EDIII-based tetravalent antigen, EDIII-T, and a biotinylated version of the latter as coating antigens. The sensitivity and specificity of these assays were compared to those obtained using the PanBio Dengue IgG/IgM ELISAs.
  
  
  RESULTS
  The performance of dengue IgG and IgM indirect ELISAs, using either a physical mixture of four EDIIIs or the single chimeric EDIII-T antigen, were comparable. Coating of a biotinylated version of the tetravalent antigen on streptavidin plates enhanced sensitivity without compromising specificity.
  
  
  CONCLUSIONS
  The incorporation of the EDIIIs of the four DENV serotypes into a single chimeric antigen did not adversely affect assay outcome in indirect ELISAs. Oriented, rather than random, immobilization of the tetravalent antigen enhanced sensitivity of detection of anti-DENV antibodies with retention of 100% specificity.
authors.affiliation: International Centre for Genetic Engineering & Biotechnology, New Delhi, India.
authors.forename: Gaurav
authors.initials: G
authors.lastname: Batra
authors.forename: Satish K
authors.initials: SK
authors.lastname: Nemani
authors.forename: Poornima
authors.initials: P
authors.lastname: Tyagi
authors.forename: Sathyamangalam
authors.initials: S
authors.lastname: Swaminathan
authors.forename: Navin
authors.initials: N
authors.lastname: Khanna
mesh_terms: D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000235:genetics; D019411:Clinical Laboratory Techniques / Q000379:methods*; D003715:Dengue / Q000175:diagnosis*; D003716:Dengue Virus / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007075:Immunoglobulin M / Q000097:blood; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012680:Sensitivity and Specificity; D014759:Viral Envelope Proteins / Q000235:genetics; D014773:Virology / Q000379:methods*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins