title: Improving dengue viral antigens detection in dengue patient serum specimens using a low pH glycine buffer treatment.
abstract: OBJECTIVE
  Early diagnosis of dengue virus (DENV) infection to monitor the potential progression to hemorrhagic fever can influence the timely management of dengue-associated severe illness. Nonstructural protein 1 (NS1) antigen detection in acute serum specimens has been widely accepted as an early diagnostic assay for dengue infection; however, lower sensitivity of the NS1 antigen-capture enzyme-linked immunosorbent assay (Ag-ELISA) in secondary dengue viral infection has been reported.
  
  
  METHODS
  In this study, we developed two forms of Ag-ELISA capable of detecting E-Ag containing virion and virus-like particles, and secreted NS1 (sNS1) antigens, respectively. The temporal kinetics of viral RNA, sNS1, and E-Ag were evaluated based on the in vitro infection experiment. Meanwhile, a panel of 62 DENV-2 infected patients' sera was tested.
  
  
  RESULTS
  The sensitivity was 3.042 ng/mL and 3.840 ng/mL for sNS1 and E, respectively. The temporal kinetics of the appearance of viral RNA, E, NS1, and infectious virus in virus-infected tissue culture media suggested that viral RNAs and NS1 antigens could be detected earlier than E-Ag and infectious virus. Furthermore, a panel of 62 sera from patients infected by DENV Serotype 2 was tested. Treating clinical specimens with the dissociation buffer increased the detectable level of E from 13% to 92% and NS1 antigens from 40% to 85%.
  
  
  CONCLUSIONS
  Inclusion of a low-pH glycine buffer treatment step in the commercially available Ag-ELISA is crucial for clinical diagnosis and E-containing viral particles could be a valuable target for acute DENV diagnosis, similar to NS1 detection.
authors.affiliation: Ph.D. Program in Microbial Genomics, National Chung-Hsing University, Taichung, Taiwan.
authors.forename: Wen-Fan
authors.initials: WF
authors.lastname: Shen
authors.affiliation: Department of Veterinary Medicine, College of Veterinary Medicine, National Chung-Hsing University, Taichung, Taiwan.
authors.forename: Jedhan Ucat
authors.initials: JU
authors.lastname: Galula
authors.affiliation: Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO, USA.
authors.forename: Gwong-Jen J
authors.initials: GJ
authors.lastname: Chang
authors.affiliation: Institute of Cellular and Organismic Biology, Academia Sinica, Taipei, 115, Taiwan.
authors.forename: Han-Chung
authors.initials: HC
authors.lastname: Wu
authors.affiliation: Institute of Epidemiology, School of Public Health, National Taiwan University, Taipei, Taiwan.
authors.forename: Chwan-Chuen
authors.initials: CC
authors.lastname: King
authors.affiliation: Graduate Institute of Microbiology and Public Health, College of Veterinary Medicine, National Chung-Hsing University, Taichung, Taiwan. Electronic address: dychao@nchu.edu.tw.
authors.forename: Day-Yu
authors.initials: DY
authors.lastname: Chao
mesh_terms: D000330:Aedes / Q000821:virology; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000936:Antigen-Antibody Complex / Q000737:chemistry; D000956:Antigens, Viral / Q000097:blood* / Q000276:immunology / Q000302:isolation & purification*; D002522:Chlorocebus aethiops; D003715:Dengue / Q000097:blood* / Q000276:immunology* / Q000821:virology; D003716:Dengue Virus / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005998:Glycine / Q000737:chemistry*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D007075:Immunoglobulin M / Q000097:blood / Q000276:immunology; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000276:immunology / Q000302:isolation & purification; D014771:Virion / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D000936:Antigen-Antibody Complex; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; C079340:NS1 protein, Dengue virus type 2; D017361:Viral Nonstructural Proteins; D005998:Glycine
keywords: Antigen-capture ELISA; Dengue; NS1 antigen; Viral particle